Method for producing polymer compound

Abstract

One aspect of the present disclosure provides a polymer compound production method comprising heating to cause a reaction between, in dimethyl sulfoxide in the presence of a base and a reducing agent, a polypeptide having at least one mercapto group, and at least one compound that has two structures represented by general formula (1) and that is selected from the group consisting of polyethers, polyesters, and polycarbonates. [In general formula (1), M represents any one of H, Na, K, NHEt3, or NHEtiPr2.]

Description

[Technical field]

[0001] The present disclosure relates to a method for producing a polymer compound, a method for producing a polymer compound solution, a method for producing a film, and a method for producing a fiber, as well as the film and the fiber. [Background technology]

[0002] Structural proteins such as silk fibroin and spider silk fibroin can exhibit excellent robustness, and therefore have been attracting attention as alternatives to structural materials composed of synthetic resins, etc. In recent years, with the advancement of recombinant technology, progress has been made in the development of mass production technology for recombinant structural proteins that mimic the above-mentioned structural proteins (e.g., Patent Document 1).

[0003] Proteins have many functional groups capable of hydrogen bonding, such as amide groups in the main chain and carboxyl, amino, hydroxyl, and mercapto groups in the side chain, and therefore many hydrogen bonds are formed between molecules. For this reason, molded bodies obtained by heating and pressurizing structural proteins are thought to have excellent mechanical strength, but they also have a fragile side, such as cracking and breaking when subjected to impacts. The above-mentioned circumstances are obstacles to using proteins as a substitute for general-purpose plastics.

[0004] As a means of modifying proteins, a method of binding a compound having properties different from those of the protein to the protein has been considered. For example, Patent Document 2 discloses a method for producing a protein material, which includes a step of binding a substance having excellent water resistance to a specific modified fibroin.

[0005] Patent Document 3 discloses a method for producing a polymer, in which a thioether bond is generated by utilizing a 1,4-addition reaction (Michael addition reaction) between a mercapto group of cysteine ​​constituting the polypeptide backbone and a carbon-carbon double bond of maleimide or a maleic acid derivative, thereby introducing at least one functional group selected from the group consisting of a polyether group, a polyester group, a polycarbonate group, a polyamide group, a polyol group, and a modified polysaccharide group into the polypeptide backbone. [Prior art documents] [Patent documents]

[0006] [Patent Document 1] International Publication No. 2015 / 178466 [Patent Document 2] International Publication No. 2019 / 194146 [Patent Document 3] International Publication No. 2021 / 187502 [Patent Document 4] Japanese Patent Application Publication No. 05-336963 [Patent Document 5] International Publication No. 1994 / 023021 [Patent Document 6] International Publication No. 2019 / 194263 Summary of the Invention [Problem to be solved by the invention]

[0007] The production method described in Patent Document 3 is useful in that various functional groups can be introduced into a polypeptide skeleton; however, depending on the structure of a Michael addition acceptor that can undergo a Michael addition reaction with a mercapto group possessed by the polypeptide skeleton, it may be necessary to more strictly control the reaction, reaction efficiency, and the like, and there is therefore room for improvement as a synthesis technique for raw materials for industrial materials.

[0008] For example, when a maleimide group is used as a Michael addition acceptor, the high reactivity of the compound serving as the Michael addition acceptor may result in the production of not only the designed molecule but also polymer compounds having unexpected long chain growth or branching, which may lead to an increase in the viscosity of the reaction solution, making it difficult to control the reaction system. In such a situation, the rate at which the raw material polypeptide becomes the desired polymer compound is low, and the raw material conversion rate tends to decrease. Furthermore, it is difficult to sufficiently control the reaction system by adjusting reaction conditions such as the acidity of the reaction solution, the reaction temperature, and the stirring speed.

[0009] Therefore, it is possible to use a Michael addition acceptor having a lower reactivity as the reaction substrate. However, this may result in a decrease in the raw material conversion rate due to the decrease in the reactivity of the substrate itself.

[0010] An object of the present disclosure is to provide a production method capable of increasing the raw material conversion rate as compared to conventional methods in the production of the above-mentioned polymer compounds. [Means for solving the problem]

[0011] According to the study by the present inventors, when the conventional manufacturing method is used on an industrial scale, when a reaction for introducing a specific functional group into a polypeptide is carried out using dimethyl sulfoxide, an aprotic polar solvent, as a solvent, the formation of a disulfide bond between mercapto groups of the polypeptide is promoted by heating during synthesis, and it is found that it may be difficult to bind the functional group to the polypeptide by a Michael addition reaction using the mercapto group. Although it is presumed that there is no big problem when the rate of the Michael addition reaction is sufficiently faster than the reaction for forming a disulfide bond, in reality, it has been confirmed that, for example, in the reaction between a substrate having a (meth)acryloyloxy group and a polypeptide, the rate of the Michael addition reaction is slow, and the reaction for forming a disulfide bond between polypeptides is promoted, so that the synthesis of the target polymer compound does not proceed sufficiently. The present disclosure has been made based on the above-mentioned findings.

[0012] This disclosure provides the following [1] to

[34] .

[0013] [1] A polypeptide having at least one mercapto group; At least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, having two structures represented by the following general formula (1): [ka] [In the general formula (1), M represents any one of H, Na, K, NHEt3, and NHEtiPr2] in dimethyl sulfoxide in the presence of a base and a reducing agent, [2] A polypeptide having at least one mercapto group; At least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, having two structures represented by the following general formula (2): [ka] [In general formula (2), R 1 represents a hydrogen atom or a methyl group. in dimethyl sulfoxide in the presence of a base, a reducing agent and a polymerization inhibitor, [3] A polypeptide having at least one mercapto group; At least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, having two structures represented by the following general formula (1): [ka] [In the general formula (1), M represents any one of H, Na, K, NHEt3, and NHEtiPr2] in dimethyl sulfoxide in the presence of a reducing agent, by heating and reacting the above. [4] A polypeptide having at least one mercapto group; At least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, having two structures represented by the following general formula (2): [ka] [In general formula (2), R 1 represents a hydrogen atom or a methyl group. in dimethyl sulfoxide in the presence of a reducing agent and a polymerization inhibitor, [5] The method for producing a polymer compound according to any one of [1] to [4], wherein the reducing agent is at least one selected from the group consisting of dithiol, sodium sulfate, sodium sulfite, and sodium dithionite. [6] The method according to any one of [1] to [5], wherein the amount of the reducing agent is 0.5 equivalents or more relative to the mercapto group of the polypeptide. [7] The method according to any one of [1] to [6], wherein the reaction between the polypeptide and the compound is carried out at 50° C. or higher. [8] A method for producing a polymer solution, comprising isolating a polymer obtained by the production method according to any one of [1] to [7] and then dissolving the isolated polymer in a solvent. [9] A method for producing a film, comprising forming a film from a polymer compound solution obtained by the production method according to [8].

[10] A method for producing fibers, comprising spinning a polymer solution obtained by the production method described in [8].

[11] A method for producing a polymer compound solution by reacting a polypeptide having at least one mercapto group with at least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, the compound having two structures represented by the following general formula (1), in dimethyl sulfoxide in the presence of a base and a reducing agent under heating; and forming a film from the polymer compound solution. [ka] [In general formula (1), M represents any one of H, Na, K, NHEt3, and NHEtiPr2.]

[12] A method for producing a polymer compound solution by reacting a polypeptide having at least one mercapto group with at least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, the compound having two structures represented by the following general formula (2), in dimethyl sulfoxide in the presence of a base, a reducing agent, and a polymerization inhibitor, under heating; and forming a film from the polymer compound solution. [ka] [In general formula (2), R 1 represents a hydrogen atom or a methyl group.

[13] A method for producing a polymer compound solution by reacting a polypeptide having at least one mercapto group with at least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, the compound having two structures represented by the following general formula (1), in dimethyl sulfoxide in the presence of a base and a reducing agent under heating; and spinning the polymer solution. [ka] [In general formula (1), M represents any one of H, Na, K, NHEt3, and NHEtiPr2.]

[14] A method for producing a polymer compound solution by reacting a polypeptide having at least one mercapto group with at least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, the compound having two structures represented by the following general formula (2), in dimethyl sulfoxide in the presence of a base, a reducing agent, and a polymerization inhibitor, under heating; and spinning the polymer solution. [ka] [In general formula (2), R 1 represents a hydrogen atom or a methyl group.

[15] A film comprising a polymer compound, The polymer compound is A polypeptide portion; and at least one structural unit selected from the group consisting of a polyether structure, a polyester structure, and a polycarbonate structure, wherein the polypeptide portion is directly bound to S in the following general formula (3), (4), or (5) via a structure represented by the following general formula (3), (4), or (5): [ka] [ka] [ka] [In the general formula (3) and the general formula (4), M represents any one of H, Na, K, NHEt3, and NHEtiPr2; in the general formula (5), R 1 indicates H or Me] A film having an elongation of 400% or more and a breaking strength greater than its yield strength.

[16] A fiber comprising a polymer compound, The polymer compound is A polypeptide portion; and at least one structural unit selected from the group consisting of a polyether structure, a polyester structure, and a polycarbonate structure, wherein the polypeptide portion is directly bound to S in the following general formula (3), (4), or (5) via a structure represented by the following general formula (3), (4), or (5): [ka] [ka] [ka] [In the general formula (3) and the general formula (4), M represents any one of H, Na, K, NHEt3, and NHEtiPr2; in the general formula (5), R 1 indicates H or Me] A fiber having an elongation of 400% or more and a breaking strength greater than its yield strength.

[17] A fiber comprising a synthetic polymer obtained by the manufacturing method according to any one of [1] to [8], the fiber having an elongation of 400% or more and a ratio of breaking strength to yield strength of more than 1.

[18] A film comprising a synthetic polymer obtained by the production method according to any one of [1] to [8], having an elongation of 400% or more and a ratio of breaking strength to yield strength of more than 1.

[19] A method for producing a solution adhesive, comprising the step of dissolving a polymer compound obtained by the method according to any one of [1] to [8] in a solvent.

[20] A method for producing a water-dispersible adhesive, comprising a step of dispersing a polymer compound obtained by the method according to any one of [1] to [8] in an aqueous medium. [twenty one] A method for producing a film-like adhesive, comprising the step of forming a film from a solution in which a polymer compound obtained by the method according to any one of [1] to [8] is dissolved. [twenty two] A method for producing a powdery adhesive, comprising the step of obtaining a powder composition containing a polymer compound obtained by the method according to any one of [1] to [8]. [twenty three] A method for producing an adherend, comprising: placing a solution in which a polymer compound obtained by the method according to any one of [1] to [8] is dissolved in a solvent between a plurality of adherends; removing the solvent from the solution; and solidifying the polymer compound, thereby adhering the adherends to each other. [twenty four] A method for producing an adherend, comprising: placing an aqueous dispersion in which a polymer compound obtained by the method according to any one of [1] to [8] is dispersed in an aqueous medium between a plurality of adherends; removing the aqueous medium from the aqueous dispersion; and solidifying the polymer compound, thereby adhering the adherends to each other. [twenty five] A method for producing an adhesive, comprising the steps of: softening a film containing a polymer compound obtained by the method according to any one of [1] to [8] by swelling or heating; interposing the film between a plurality of adherends; and curing the film while the film is pressed against the adherends, thereby bonding the adherends together.

[26] A method for producing an adherend, comprising: heating a powder composition containing a polymer compound obtained by the method according to any one of [1] to [8] between a plurality of adherends; and applying pressure to the powder composition through the adherends to solidify the powder composition, thereby adhering the adherends to each other.

[27] A method for producing a solution-type coating agent, comprising a step of dissolving a polymer compound obtained by the method according to any one of [1] to [8] in a solvent.

[28] A method for producing a water-dispersible coating agent, comprising a step of dispersing a polymer compound obtained by the method according to any one of [1] to [8] in an aqueous medium.

[29] A method for producing a film-like coating agent, comprising a step of forming a film from a solution in which a polymer compound obtained by the method according to any one of [1] to [8] is dissolved.

[30] A method for producing a powder coating agent, comprising a step of obtaining a powder composition containing a polymer compound obtained by the method according to any one of [1] to [8].

[31] A method for producing a laminate having a substrate and a coating layer formed on at least a portion of a surface of the substrate, the method comprising the steps of: A method for producing a laminate, comprising: supplying a solution in which a polymer compound obtained by the method according to any one of [1] to [8] is dissolved in a solvent to at least a part of a surface of the substrate; coating at least a part of the surface of the substrate with the solution; and removing the solvent from the solution to solidify the polymer compound, thereby forming a coating layer on at least a part of the surface of the substrate.

[32] A method for producing a laminate having a substrate and a coating layer formed on at least a portion of a surface of the substrate, the method comprising the steps of: A method for producing a laminate, comprising: supplying an aqueous dispersion in which a polymer compound obtained by the method according to any one of [1] to [8] is dispersed in an aqueous medium to at least a part of a surface of a substrate; coating at least a part of the surface of the substrate with the aqueous dispersion; removing the aqueous medium from the aqueous dispersion; and solidifying the polymer compound, thereby forming a coating layer on at least a part of the surface of the substrate.

[33] A method for producing a laminate having a substrate and a coating layer formed on at least a portion of a surface of the substrate, the method comprising the steps of: A method for producing a laminate, comprising the steps of: softening a film containing a polymer compound obtained by the method according to any one of [1] to [8] by swelling or heating; placing the film on at least a part of a surface of the substrate; and curing the film in a state of being pressed against the substrate, thereby forming the coating layer on at least a part of the surface of the substrate.

[34] A method for producing a laminate having a substrate and a coating layer formed on at least a portion of a surface of the substrate, the method comprising the steps of: A method for producing a laminate, comprising: placing a powder composition containing a polymer compound obtained by the method according to any one of [1] to [8] on at least a part of a surface of a substrate; heating the powder composition while pressing the powder composition between a pressurizing body and the substrate to solidify the powder composition, thereby forming the coating layer on at least a part of the surface of the substrate.

[0014] One aspect of the present disclosure provides a method for producing a polymer compound, comprising heating and reacting a polypeptide having at least one mercapto group with at least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, the compound having two structures represented by the following general formula (1), in dimethyl sulfoxide in the presence of a base and a reducing agent: [ka] [In the general formula (1), M represents any one of H, Na, K, NHEt3, and NHEtiPr2]

[0015] In one example of the above-mentioned production method, a method is adopted in which a base and a reducing agent are present in the reaction system when the polypeptide is reacted with a predetermined compound disclosed herein. The reducing agent is stable in dimethyl sulfoxide, does not easily cause a Michael addition reaction with the functional group represented by formula (1), and can inhibit the formation of disulfide bonds between mercapto groups of the polypeptide. The base can activate the mercapto group of the polypeptide and promote the Michael addition reaction with the functional group represented by formula (1). Due to this action, a skeleton such as a polyether can be introduced into the polypeptide by the method of heating in dimethyl sulfoxide, and a polymer compound can be produced with higher efficiency. When the production conditions are the same as the other conditions, the base can be blended to obtain the desired effect of the present disclosure at a higher level, but it can be omitted depending on the level of the desired effect.

[0016] One aspect of the present disclosure provides a method for producing a polymer compound, comprising heating and reacting a polypeptide having at least one mercapto group with at least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, the compound having two structures represented by the following general formula (2), in dimethyl sulfoxide in the presence of a base, a reducing agent, and a polymerization inhibitor: [ka] [In general formula (2), R 1 represents a hydrogen atom or a methyl group.

[0017] In the above-mentioned example of the production method, a method is adopted in which a base and a reducing agent are present in the reaction system when the polypeptide is reacted with a predetermined compound of the present disclosure. Here, the functional group represented by the general formula (2) is radically polymerized by heating in dimethyl sulfoxide, and self-polymerization of at least one compound selected from the group consisting of polyether, polyester, and polycarbonate having two structures represented by the general formula (2) may occur. However, according to the production method of the present disclosure, the radical polymerization reaction of the functional group represented by the general formula (2) can be suppressed by also having a polymerization inhibitor present in the reaction system. Due to this action, even by the method of heating in dimethyl sulfoxide, a skeleton such as polyether can be introduced into the polypeptide, and a polymer compound can be produced with higher efficiency. When the production conditions are the same as the other conditions, the base can be blended to obtain the desired effect of the present disclosure at a higher level, but it can be omitted depending on the level of the desired effect.

[0018] In the above-mentioned production method, the reducing agent may be at least one selected from the group consisting of dithiol, sodium sulfate, sodium sulfite, and sodium dithionite. Such a specific reducing agent is highly stable in dimethyl sulfoxide and can further inhibit the formation of disulfide bonds between mercapto groups in the polypeptide.

[0019] In the above-mentioned production method, the amount of the reducing agent may be 0.5 equivalents or more relative to the mercapto groups of the polypeptide. By setting the amount of the reducing agent within the above range, the formation of disulfide bonds caused by the reaction between the mercapto groups of the polypeptide can be more sufficiently inhibited. This can further improve the production efficiency of the target synthetic polymer.

[0020] In the above production method, the reaction between the polypeptide and the compound may be carried out at 50° C. or higher.

[0021] One aspect of the present disclosure provides a method for producing a polymer solution, the method comprising dissolving the polymer obtained by the above-mentioned production method in a solvent.

[0022] A method for producing a polymer film, comprising forming a film from the polymer solution obtained by the above-mentioned method for producing a polymer solution. The forming into the film may be, for example, cast molding.

[0023] One aspect of the present disclosure provides a method for producing a polymer fiber, comprising spinning a polymer compound solution obtained by the above-described production method.

[0024] One aspect of the present disclosure provides a film comprising a polymer compound, the polymer compound having a structure in which a polypeptide portion and at least one structural unit selected from the group consisting of a polyether structure, a polyester structure, and a polycarbonate structure are directly bonded to each other via a structure represented by the following formula (3), (4), or (5) such that the polypeptide portion is bonded to S in the following formula (3), (4), or (5), the film having an elongation of 400% or more and a breaking strength greater than a yield strength. In formulas (3) and (4), M represents H, Na, K, NHEt3, or NHEtiPr2, and in formula (5), R1 represents H or Me.

[0025] [ka]

[0026] [ka]

[0027] [ka]

[0028] One aspect of the present disclosure provides a fiber comprising a polymer compound, or a film comprising the polymer compound, the polymer compound having a structure in which a polypeptide portion and at least one structural unit selected from the group consisting of a polyether structure, a polyester structure, and a polycarbonate structure are directly bonded to each other via a structure represented by the following general formula (3), (4), or (5) such that the polypeptide portion is bonded to S in the following general formula (3), (4), or (5), the fiber having an elongation of 400% or more and a breaking strength greater than a yield strength. In general formulas (3) and (4), M represents H, Na, K, NHEt3, or NHEtiPr2, and in general formula (5), R1 represents H or Me.

[0029] [ka]

[0030] [ka]

[0031] [ka] Effect of the Invention

[0032] According to the present disclosure, in the production of the above-mentioned polymer compound, a production method capable of increasing the raw material conversion rate compared to conventional methods can be provided. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0033] Unless otherwise specified, the materials exemplified in this specification can be used alone or in combination of two or more. When a plurality of substances corresponding to each component are present in the composition, the content of each component in the composition means the total amount of the plurality of substances present in the composition, unless otherwise specified.

[0034] In this specification, the molding material refers to a material used to produce a molded body. The molding material according to the present disclosure can also be said to be a material obtained by chemically modifying a polypeptide. The shape of the molded body in this specification is not particularly limited, and may be, for example, a film shape, a plate shape, a block shape, a sponge shape, a fiber shape, etc. The form of the molding material in this specification is also not limited in any way, and may be, for example, a powder, a granule, a liquid, a gel, etc. In addition, the molding material according to the present disclosure can be made into a molded body of various shapes by, for example, hot pressure molding, cast molding, spinning, etc. When molding, a mold may be used as necessary.

[0035] The method for producing a polymer compound according to the present disclosure utilizes a reaction between a mercapto group (nucleophilic functional group) of a polypeptide and a skeleton such as a polyether having a specific unsaturated bond (electrophilic functional group).

[0036] The first embodiment of the method for producing a polymer compound comprises reacting a polypeptide having at least one mercapto group (hereinafter also referred to as component (A)) with at least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, having two structures represented by the following formula (1) (hereinafter also referred to as component (B1) or, together with component (B2) described below, simply referred to as component (B)), by heating in dimethyl sulfoxide in the presence of a base (hereinafter also referred to as component (C)) and a reducing agent (hereinafter also referred to as component (D)). The heating reaction is a reaction between component (A) and component (B), and is carried out, for example, by heating to 50°C or higher. The timing of heating is not limited to after mixing of components (A) to (D), and for example, components (A) to (D) may be added to preheated dimethyl sulfoxide, components (C) and (D) may be added to dimethyl sulfoxide, heating may be started, and after a predetermined temperature is reached, components (A) and (B) may be added and reacted, or one of components (A) and (B) may be added to components (C) and (D), and heating may be started, and after a predetermined temperature is reached, the remaining one of components (A) and (B) may be added and reacted. When the manufacturing conditions are the same for all other conditions, the incorporation of component (C) allows the desired effect of the present disclosure to be obtained at a higher level, but it may be omitted depending on the level of the desired effect.

[0037] [ka]

[0038] The number of mercapto groups possessed by the above polypeptide (component (A)) is 1 or more per molecule, and may be, for example, 2 or more, 4 or more, 8 or more, or 16 or more. When the number of the mercapto groups is within the above range, the contact probability of the reaction sites is improved, and the reaction can proceed efficiently. The number of mercapto groups possessed by the component (A) per molecule may be, for example, 64 or less, or 32 or less. When the number of the mercapto groups is within the above range, the production of by-products formed by the reaction between the mercapto groups possessed by the polypeptide can be suppressed, and the target polymer compound can be obtained more efficiently.

[0039] The number of amino acid residues constituting the (A) component may be, for example, 50 or more, 100 or more, 150 or more, 200 or more, 250 or more, 300 or more, 350 or more, 400 or more, 450 or more, or 500 or more. The number of amino acid residues may be, for example, 5000 or less, 4500 or less, 4000 or less, 3500 or less, 3000 or less, 2500 or less, 2000 or less, 1500 or less, or 1000 or less. The smaller the number of amino acid residues, the higher the solubility in dimethyl sulfoxide tends to be. When the number of amino acid residues constituting the (A) component is, for example, 5000 or less or 2500 or less, the production efficiency of the target polymer compound can be further improved when the polypeptide is dissolved in a solvent and reacted with the above compound (component (B1)). In addition, the molded article composed of the obtained polymer compound can exhibit a higher level of mechanical strength and flexibility.

[0040] The (A) component may be, for example, a hydrophobic polypeptide. When the (A) component is a hydrophobic polypeptide, the (A) component can be prepared more easily, and the raw material cost in the above-mentioned production method can be further reduced. In this case, the molded article made of the obtained polymer compound can exhibit better flexibility. The molded article can also exhibit better water resistance. In this case, the obtained molded article can have a longer service life. The hydrophobicity of the (A) component can be adjusted as desired to that of the polymer compound.

[0041] The hydrophobicity of component (A) can be estimated using the value of the average hydropathy index (hereinafter, sometimes referred to as average HI) as an index. The average HI value of component (A) may be more than 0, for example, 0.10 or more, 0.20 or more, 0.22 or more, 0.25 or more, 0.30 or more, 0.35 or more, 0.40 or more, 0.45 or more, 0.50 or more, 0.55 or more, 0.60 or more, 0.65 or more, or 0.70 or more. The upper limit of the average HI value of component (A) is not particularly limited, but may be, for example, 1.00 or less, or 0.7 or less.

[0042] The component (A) is preferably one that has low solubility in an aqueous lithium bromide solution (concentration: 9M) at 60°C. This solubility can be evaluated using a polypeptide corresponding to the component (A). The maximum concentration of the polypeptide when dissolved in an aqueous lithium bromide solution (concentration: 9M) at 60°C may be, for example, less than 30% by mass, less than 25% by mass, less than 20% by mass, less than 15% by mass, less than 10% by mass, less than 5% by mass, or less than 1% by mass. The component (A) may be one that is completely insoluble in an aqueous lithium bromide solution (concentration: 9M) at 60°C. The solubility of the polypeptide portion of the polymer compound obtained by the production method according to the present disclosure can be confirmed by using a polypeptide obtained by decomposing the polymer compound and isolating only the hydrophobic polypeptide.

[0043] It is preferable that the component (A) has a large contact angle with water. The contact angle with water can be evaluated by using a polypeptide corresponding to the component (A) on a substrate. More specifically, a film made of the above polypeptide is formed and evaluated by using the film. A polypeptide constituting a film in which the contact angle is 55° or more after 5 seconds has elapsed after water is dropped on the film is preferable as the component (A). The contact angle may be, for example, 60° or more, 65° or more, or 70° or more. For the polymer compound obtained by the production method according to the present disclosure, the contact angle of the polypeptide portion of the polymer compound can be confirmed by using a polypeptide obtained by decomposing the polymer compound and isolating only the hydrophobic polypeptide.

[0044] The component (A) is preferably excellent in hot water resistance. The hot water resistance can be evaluated using a polypeptide corresponding to the component (A). A dispersion liquid consisting of the above polypeptide and water is prepared, and the content of the above polypeptide is 5% by mass. The component (A) is preferably a polypeptide that does not decompose even when the dispersion is heated at 100°C for 5 hours. The above hot water resistance of the polypeptide portion of the polymer compound obtained by the production method according to the present disclosure can be confirmed by using a polypeptide obtained by decomposing the polymer compound and isolating only the hydrophobic polypeptide.

[0045] The weight average molecular weight of the (A) component is, for example, 200 to 1,000,000, more preferably 300 to 900,000, even more preferably 400 to 800,000, even more preferably 500 to 700,000, even more preferably 600 to 600,000, even more preferably 1,000 to 600,000, even more preferably 3,000 to 600,000, even more preferably 5,000 to 600,000, even more preferably 10,000 to 600,000, and even more preferably 5,000 to 100,000. If the weight average molecular weight of the (A) component is less than 200, the polypeptide that functions as a hard segment may be too small relative to the polyether structure (soft segment) of the (B1) component. In this case, the rigidity of the molded body molded using the obtained polymer compound (molding material) may be reduced, making it difficult to use it as a structure, for example.

[0046] The weight-average molecular weight in this specification is a value measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The electrophoresis is carried out according to the following procedure. That is, first, 200 μL of 2M lithium chloride DMSO (manufactured by Fujifilm Wako Pure Chemical Industries, Ltd.) is added to 2 mg of powder sample, and the sample is dissolved by stirring while heating at 80° C. for 60 minutes and then at 95° C. for 10 minutes. Thereafter, the sample is diluted 50 times with 10M urea solution, further diluted 2 times with sample buffer (manufactured by Fujifilm Wako Pure Chemical Industries, Ltd.), and heated at 95° C. for 5 minutes to denature the protein. Next, an SDS-PAGE gel (manufactured by Bio-lad) is attached to an electrophoresis device (manufactured by Bio-lad), and the device is filled with SDS buffer, while the electrophoresis device is connected to a power supply device (manufactured by Bio craft). 10 μL of the denatured sample was added to each well of the SDS-PAGE gel, and a current of 30 mA was applied for 30 minutes. After electrophoresis, the SDS-PAGE gel was removed from the device, immersed in Oriole fluorescent gel stain (Bio-lad), and shaken for 1 hour. The gel was then placed on a UV sample tray (Bio-lad) and the stained image was captured using a Gel Doc EZ gel camera (Bio-lad). XL ladder Broad (Apro Science) was used as the standard sample.

[0047] The molecular weight of the (A) component may be, for example, 2 kDa or more, 3 kDa or more, 4 kDa or more, 5 kDa or more, 6 kDa or more, 7 kDa or more, 8 kDa or more, 9 kDa or more, 10 kDa or more, 20 kDa or more, 30 kDa or more, 40 kDa or more, 50 kDa or more, 60 kDa or more, 70 kDa or more, 80 kDa or more, 90 kDa or more, or 100 kDa or more. The molecular weight of the (A) component may be, for example, 500 kDa or less, 400 kDa or less, less than 360 kDa, 300 kDa or less, or 200 kDa or less. The molecular weight of the (A) component may be adjusted within the above-mentioned range, and may be, for example, 2 to 500 kDa. The molecular weight can be measured by a liquid chromatograph mass spectrometer.

[0048] The component (A) may be a natural protein or an artificial protein. The component (A) may also be a protein (modified protein) obtained by chemically modifying a natural protein or an artificial protein (e.g., alkylation, etherification, esterification, amidation, thioetherification, thioesterification, etc.). The amino acid sequence of the natural protein, artificial protein, or modified protein is not particularly limited as long as it has one or more mercapto groups.

[0049] As the component (A), for example, a protein having similar physical properties can be adopted in accordance with the required characteristics according to the application of the polymer compound. Examples of the polypeptide according to the present embodiment include a polypeptide that can be used for medical purposes and a polypeptide that can be used for industrial purposes. "Usable for industrial purposes" means that it can be used, for example, as various general-purpose materials used indoors and outdoors.

[0050] Examples of polypeptides that can be used for medical purposes include enzymes, regulatory proteins, receptors, peptide hormones, cytokines, membrane or transport proteins, antigens used in vaccination, vaccines, antigen-binding proteins, immunostimulating proteins, allergens, full-length antibodies or antibody fragments or derivatives.

[0051] Specific examples of polypeptides that can be used industrially include structural proteins. Structural proteins refer to proteins involved in the structure of a living organism, or proteins that constitute structures created by a living organism, or proteins derived therefrom. Structural proteins also refer to proteins that self-aggregate under certain conditions to form structures such as fibers, films, resins, gels, micelles, and nanoparticles. Furthermore, structural proteins can be said to be proteins that have repeated motifs consisting of characteristic amino acid sequences or amino acid residues and form the skeletons of living organisms and materials. Specific examples of structural proteins include spider silk, silkworm silk, keratin, collagen, elastin, and resilin, as well as proteins derived therefrom. Structural proteins may be artificial fibroins or artificial spider silk fibroins (artificially modified spider silk fibroins).

[0052] The structural protein may be an artificial structural protein. Artificial structural proteins include synthetic proteins and recombinant structural proteins produced from microorganisms by genetic recombination technology. That is, in this specification, "artificial structural protein" means an artificially produced structural protein. The artificial structural protein may have the same amino acid sequence as a natural structural protein, or may be a modified structural protein in which a part of the amino acid sequence is modified based on the amino acid sequence of a naturally-occurring structural protein from the standpoint of productivity, moldability, etc.

[0053] The artificial structural protein may have a glycine residue content of 10 to 55%. The glycine residue content may be, for example, 13% to 55%, 15% to 55%, 18% to 55%, 20% to 55%, 22% to 55%, or 25% to 55%. In this specification, the "glycine residue content" is a value represented by the following formula.

[0054] Glycine residue content = (number of glycine residues contained in artificial structural protein / total number of amino acid residues in polypeptide) x 100 (%)

[0055] The artificial structural protein may have 150 or more amino acid residues. The number of amino acid residues may be, for example, 200 or more or 250 or more, and is preferably 300 or more, 350 or more, 400 or more, 450 or more, or 500 or more.

[0056] The artificial structure protein may have a total content (total content) of at least one amino acid residue selected from the group consisting of serine, threonine, and tyrosine (i.e., any of the serine residue content, the threonine residue content, the tyrosine residue content, the sum of the serine residue content and the threonine residue content, the sum of the serine residue content and the tyrosine residue content, the sum of the threonine residue content and the tyrosine residue content, the sum of the serine residue content, the threonine residue content and the tyrosine residue content), the alanine residue content, and the glycine residue content) of 40% or more. The total content may be, for example, 45% or more, 50% or more, 55% or more, or 60% or more. There is no particular upper limit to the total content, but it may be, for example, 90% or less, 85% or less, or 80% or less.

[0057] The total of the serine residue content, threonine residue content and tyrosine residue content of the artificial structural protein may be 4% or more, 4.5% or more, 5% or more, 5.5% or more, 6% or more, 6.5% or more, or 7% or more. The total of the serine residue content, threonine residue content and tyrosine residue content may be, for example, 35% or less, 33% or less, 30% or less, 25% or less, or 20% or less.

[0058] The artificial structural protein has an average distribution of serine residues, threonine residues, or tyrosine residues, and the total content of serine residues, threonine residues, and tyrosine residues in any 20 consecutive amino acid residues may be 4% or more, 5% or more, 10% or more, or 15% or more, or may be 50% or less, 40% or less, 30% or less, or 20% or less.

[0059] The alanine residue content, serine residue content, threonine residue content and tyrosine residue content have the same meaning as those obtained by replacing the alanine residue with a glycine residue, serine residue, threonine residue and tyrosine residue, respectively, in the above formula.

[0060] The artificial structural protein may have a repeat sequence. That is, the artificial structural protein may have a plurality of amino acid sequences (repeat sequence units) with high sequence identity within the artificial structural protein. The number of amino acid residues in the repeat sequence unit is preferably 6 to 200.

[0061] The total number of glycine residues, serine residues, glutamine residues and alanine residues relative to the total number of amino acid residues in the repeat sequence unit may be 40% or more, 45% or more, 50% or more, 55% or more, 60% or more, 65% or more, or 70% or more. The sequence identity between the repeat sequence units may be, for example, 80% or more, 85% or more, 90% or more, 95% or more, 96% or more, 97% or more, 98% or more, or 99% or more.

[0062] The hydrophobicity index of the repeating sequence unit may be, for example, -0.80 or more, -0.70 or more, -0.60 or more, -0.50 or more, -0.40 or more, -0.30 or more, -0.20 or more, -0.10 or more, 0.00 or more, 0.22 or more, 0.25 or more, 0.30 or more, 0.35 or more, 0.40 or more, 0.45 or more, 0.50 or more, 0.55 or more, 0.60 or more, 0.65 or more, or 0.70 or more. The upper limit of the hydrophobicity index of the repeating sequence unit is not particularly limited, but may be, for example, 1.0 or less, or 0.7 or less.

[0063] The artificial structural protein may contain an (A)n motif. In the present specification, the (A)n motif means an amino acid sequence mainly composed of alanine residues. The number of amino acid residues in the (A)n motif may be 2 to 27, and may be an integer of 2 to 20, 2 to 16, or 2 to 12.

[0064] In addition, the ratio of the number of alanine residues to the total number of amino acid residues in the (A)n motif may be 40% or more, and may be 45% or more, 50% or more, 55% or more, 60% or more, 65% or more, 70% or more, 80% or more, 83% or more, 85% or more, 86% or more, 90% or more, 95% or more, or 100% (meaning that it is composed only of alanine residues).

[0065] The (A)n motif may be 80% or more in terms of the total number of alanine residues, serine residues, threonine residues, and valine residues relative to the total number of amino acid residues in the (A)n motif, but is preferably 85% or more, more preferably 90% or more, even more preferably 95% or more, and even more preferably 100% (meaning that it is composed of only one or more amino acid residues selected from alanine residues, serine residues, threonine residues, and valine residues). The (A)n motifs present in the recombinant structural protein according to the present embodiment may have the same amino acid sequence or different amino acid sequences. Since the (A)n motif mainly contains alanine residues, it is likely to form an α-helix structure or a β-sheet structure. Since the (A)n motif is included in the repeating sequence unit, the artificial structural protein according to the present embodiment will have these secondary structures repeatedly. Therefore, when the artificial structural protein is formed into a molded product such as a fiber, film, or resin, as described below, it is expected that the secondary structures will provide high strength.

[0066] In addition, when forming a structural protein, an amino acid having a relatively small side chain is more likely to form a hydrogen bond, and a molded product having higher strength is more likely to be obtained. In addition, since the side chains of alanine residues and glycine residues are non-polar amino acids, they are arranged so as to face inward during the folding process in the production of a polypeptide, and tend to form an α-helix structure or a β-sheet structure. Therefore, it is desirable to have a high ratio of amino acids such as glycine residues and alanine residues. From the viewpoint of obtaining a molded product having superior strength, the alanine residue content may be, for example, 10 to 40%, and may be 12 to 40%, 15 to 40%, 18 to 40%, 20 to 40%, or 22 to 40%. From the viewpoint of obtaining a molded product having superior strength, the glycine residue content may be, for example, 10 to 55%, and may be 11 to 55%, 13 to 55%, 15 to 55%, 18 to 55%, 20 to 55%, 22 to 55%, or 25 to 55%.

[0067] As used herein, the "alanine residue content" refers to the number of alanine residues relative to the total number of amino acid residues constituting a protein, and is a value represented by the following formula.

[0068] Alanine residue content = (number of alanine residues in protein / total number of amino acid residues in protein) x 100 (%)

[0069] In addition, the glycine residue content, serine residue content, threonine residue content, proline residue content and tyrosine residue content have the same meaning as those obtained by replacing the alanine residue in the above formula with glycine residue, serine residue, threonine residue, proline residue and tyrosine residue, respectively.

[0070] The structural protein preferably contains amino acids with relatively large side chains or amino acids with flexibility homogeneously to a certain extent throughout the entire sequence. Specifically, the structural protein may contain motifs containing tyrosine residues, threonine residues, and proline residues in a repeated cycle. Such a structural protein is likely to inhibit the formation of strong intermolecular hydrogen bonds during processing of a molded product obtained by molding, and is likely to improve processability. For example, the total content of proline residues, threonine residues, and tyrosine residues in any 20 consecutive amino acid residues may be 5% or more, more than 5.5%, 6.0% or more, more than 6.5%, 7.0% or more, more than 7.5%, 8.0% or more, more than 8.5%, 9.0% or more, 10.0% or more, or 15.0% or more. For example, the total content of proline residues, threonine residues, and tyrosine residues in any 20 consecutive amino acid residues may be 50% or less, 40% or less, 30% or less, or 20% or less.

[0071] From the viewpoint of improving the productivity of the reaction-modified protein, the artificial structure protein may have a total content of serine residues, threonine residues and tyrosine residues of, for example, 4% or more, 4.5% or more, 5% or more, 5.5% or more, 6% or more, 6.5% or more, or 7% or more. The total content of serine residues, threonine residues and tyrosine residues may be, for example, 35% or less, 33% or less, 30% or less, 25% or less, or 20% or less.

[0072] In the above production method, the amount of the polypeptide (component (A)) may be, for example, more than 1 part by mass, 2 parts by mass or more, 5 parts by mass or more, more than 5 parts by mass, 6 parts by mass or more, 7 parts by mass or more, or 8 parts by mass or more, relative to 100 parts by mass of dimethyl sulfoxide. Since the production method according to the present disclosure can suppress the formation of disulfide bonds between polypeptides, it is possible to proceed with the production of polymer compounds in a state where the concentration of the polypeptide is increased, and more efficient production may be possible. The amount of component (A) may be, for example, 50 parts by mass or less, 30 parts by mass or less, 20 parts by mass or less, or 15 parts by mass or less, relative to 100 parts by mass of dimethyl sulfoxide. When the upper limit of the amount of component (A) is within the above range, the reaction can be progressed while more sufficiently suppressing the generation of by-products and gelation. In the above production method, the proportion of the above polypeptide (component (A)) in the dimethyl sulfoxide solution may be, for example, 15% by mass or less, less than 15% by mass, 14% by mass or less, 12.5% ​​by mass or less, or 11.5% by mass or less.

[0073] At least one compound (component (B1)) selected from the group consisting of polyether, polyester, and polycarbonate, which has two structures represented by formula (1), is a compound that, due to having a structure represented by formula (1), is capable of reacting with a mercapto group in the above-mentioned polypeptide and is capable of introducing a polyether structure, a polyester structure, or a polycarbonate structure into the polypeptide.

[0074] Examples of polyethers having two structures represented by formula (1) include polyethylene glycol, polytetramethylene glycol, polypropylene glycol, and ethylene glycol-propylene glycol copolymers. Examples of polyesters having two structures represented by formula (1) include polylactic acid, polyglycolic acid, polybutylene succinate, polycaprolactone, and polyhydroxyalkanoates including 3-hydroxybutanoic acid-3-hydroxyhexanoic acid copolymers. Examples of polycarbonates having two structures represented by formula (1) include aliphatic polycarbonates such as polyethylene carbonate, polypropylene carbonate, and polytrimethylene carbonate. These compounds have easier molecular motion than the above-mentioned polypeptides, and therefore, when they are the above-mentioned compounds, the flexibility of the molded article made of the obtained polymer compound can be further improved.

[0075] In the above production method, the weight average molecular weight of the compound (component (B1)) may be, for example, 100 or more, 1000 or more, or 2000 or more. By using a component (B1) having a weight average molecular weight within the above range, the effect of bulkiness between molecules can be further suppressed, and the reaction can proceed efficiently. In the above production method, the weight average molecular weight of component (B1) may be, for example, 200,000 or less, 100,000 or less, 50,000 or less, or 10,000 or less. By using a component (B1) having a weight average molecular weight within the above range, the decrease in the solubility of the compound in solvents and the decrease in reactivity with polypeptides can be more sufficiently suppressed.

[0076] In the above production method, the weight-average molecular weight of the compound (component (B1)) may be, for example, 0.005 times or more, 0.01 times or more, 0.05 times or more, or 0.1 times or more based on the weight-average molecular weight of the polypeptide. By setting the relationship between the weight-average molecular weight of the polypeptide and the component (B1) within the above range, it is possible to produce a synthetic polymer having a polyether structure or the like of sufficient length relative to the polypeptide portion. A molded product composed of such a synthetic polymer can have excellent flexibility. In the above production method, the weight-average molecular weight of the component (B1) may be, for example, 20 times or less, 10 times or less, 5 times or less, or 2 times or less based on the weight-average molecular weight of the polypeptide. By setting the relationship between the weight-average molecular weight of the polypeptide and the component (B) within the above range, the reaction can proceed while further suppressing the decrease in reactivity. In the above production method, the weight average molecular weight of the (B1) component may be adjusted within the above-mentioned range, and may be, for example, 0.005 to 20 times, 0.01 to 10 times, 0.05 to 5 times, or 0.1 to 2 times the weight average molecular weight of the above polypeptide.

[0077] In the above production method, the ratio of the number of moles of the structure represented by formula (1) in the above compound (component (B1)) to the number of moles of the mercapto group in the above polypeptide (component (A)) may be, for example, 0.1 times or more, 0.2 times or more, 0.5 times or more, or 0.7 times or more. By setting the above ratio within the above range, the reaction can proceed while further suppressing the decrease in reactivity. In the above production method, the ratio of the number of moles of the structure represented by formula (1) in the (B1) component to the number of moles of the mercapto group in the (A) component may be, for example, 5.0 times or less, 3.0 times or less, 1.5 times or less, or 0.8 times or less. By setting the above ratio within the above range, the reaction can proceed while further suppressing the generation of by-products and gelation. By adjusting the above molar ratio, the number of polyether structures and the like in the obtained polymer compound can be adjusted. In the obtained polymer compound, the total number of polyether structures etc. per polypeptide can be, for example, 1 or more, or 2 or more, and can be 2 to 10, 2 to 8, 2 to 6, or 2 to 4.

[0078] The number of moles of sulfhydryl groups in this specification means a value measured by enzymatically cutting out the polypeptide in the measurement sample, converting it into amino acids, and then separating and detecting them by liquid chromatography. When preparing a polypeptide by oneself, the number of moles of the above functional groups can be determined from the sequence of the designed molecule.

[0079] In the present specification, the number of moles of the structure represented by general formula (1) and the structure represented by general formula (2) refers to a value measured by MALDI-TOFMS.

[0080] In the above production method, the molar number of the compound (component (B1)) present in dimethyl sulfoxide may be, for example, 0.1 times or more, 0.2 times or more, 0.5 times or more, or 0.8 times or more relative to the total molar number of mercapto groups of the polypeptide. By adjusting the raw material blending ratio so that the molar number of the (B1) component is greater than the total molar number of mercapto groups, the probability of reaction between the polypeptide and the compound can be improved and the frequency of disulfide bond formation between the polypeptides can be more sufficiently reduced. In the above production method, the molar number of the (B1) component present in dimethyl sulfoxide may be, for example, 5.0 times or less, 3.0 times or less, 1.5 times or less, or 1.0 times or less relative to the total molar number of mercapto groups of the polypeptide. By setting the molar number of the (B1) component within the above range, the reaction can be progressed while further suppressing the generation of by-products and gelation.

[0081] The base (component (C)) in the above-mentioned production method is a compound that can activate the mercapto group of the polypeptide and promote the Michael addition reaction with the functional group represented by formula (1). Examples of the base include primary amines, secondary amines, tertiary amines, nitrogen-containing ring compounds, nitrogen-containing aromatic compounds, and inorganic bases. Examples of primary amines include ethanolamine and hexamethylenediamine. Examples of secondary amines include diethylamine, methylethylamine, and N-methylbutylamine. Examples of tertiary amines include triethylamine, diethylmethylamine, and DIPEA (N,N-diisopropylethylamine). Examples of the nitrogen-containing ring compound include quinuclidine, DABCO (1,4-diazabicyclo[2.2.2]octane), DBU (1,8-diazabicyclo[5.4.0]undec-7-ene), and DBN (1,5-diazabicyclo[4.3.0]non-5-ene). Examples of the nitrogen-containing aromatic compound include pyridine and imidazole. Examples of the inorganic base include sodium hydroxide, potassium hydroxide, sodium bicarbonate, potassium bicarbonate, sodium carbonate, potassium carbonate, cesium carbonate, tripotassium phosphate, sodium acetate, and potassium acetate.

[0082] The lower limit of the amount of the base may be, for example, 0.01 parts by mass or more, 0.05 parts by mass or more, 0.1 parts by mass or more, or 0.5 parts by mass or more, based on the mass of the polypeptide. By setting the lower limit of the amount of the base within the above range, the reaction between the mercapto group of the polypeptide and the unsaturated bond represented by formula (1) or general formula (2) of the compound can be further promoted. The upper limit of the amount of the base may be, for example, 50 parts by mass or less, 20 parts by mass or less, 10 parts by mass or less, or 5 parts by mass or less, based on the mass of the polypeptide. By setting the upper limit of the amount of the base within the above range, the decomposition of the molecule due to the influence of the remaining base can be further suppressed, and the desired polymer compound can be synthesized. The amount of the base may be adjusted within the above range, and may be, for example, 0.05 to 50 parts by mass, or 0.1 to 10 parts by mass, based on the mass of the polypeptide. The amount of the base added may be, for example, 0 to 50 parts by mass, 0 to 10 parts by mass, 0 to 1 part by mass, or 0 to 0.5 parts by mass based on the mass of the polypeptide.

[0083] The reducing agent (component (D)) in the above-mentioned production method is a compound that can stabilize the mercapto group of the polypeptide in dimethyl sulfoxide and inhibit the formation of a disulfide bond between the mercapto groups. The reducing agent may contain at least one selected from the group consisting of thiol, dithiol, sodium sulfite, sodium hyposulfite, sodium sulfate, and sodium dithionite, and the reducing agent may contain at least one selected from the group consisting of dithiol and sodium sulfite, and may be dithiol or sodium sulfite. The reducing agent is preferably a compound having a mercapto group. The reducing agent is preferably a dithiol.

[0084] The compound having a mercapto group is preferably a compound represented by the following general formula (Y). Such a compound forms a chemically stable six-membered ring structure in the oxidized form after the reduction reaction, suppressing the reaction between the reducing agent and the Michael addition acceptor in the reaction system, and also suppresses the formation of disulfide bonds between polypeptides by forming a ring structure between the reducing agent and the mercapto group of the polypeptide, making the reaction between the mercapto group of the polypeptide and the Michael addition acceptor significant, and can more efficiently produce the target polymer compound according to the reaction design. Examples of dithiols include dithiothreitol and 1,4-butanedithiol. The compound (dithiol) represented by general formula (Y) is specifically dithiothreitol or 1,4-butanedithiol.

[0085] [ka]

[0086] Other compounds having a mercapto group include, for example, 3-mercaptopropionic acid, 3-mercapto-1,2-propanediol, and pentaerythritol tetra(3-mercaptopropionate). In addition to the above compounds, the reducing agent may be, for example, tris(2-carboxyethyl)phosphine (TCEP).

[0087] The lower limit of the amount of the reducing agent may be, for example, 0.1 equivalents or more, 0.3 equivalents or more, 0.5 equivalents or more, 0.8 equivalents or more, or 1.0 equivalents or more, relative to the mercapto groups of the polypeptide. By setting the lower limit of the amount of the reducing agent within the above range, the formation of disulfide bonds due to the reaction between the mercapto groups of the polypeptide can be more sufficiently inhibited. This can further improve the production efficiency of the target synthetic polymer. The upper limit of the amount of the reducing agent may be, for example, 2.4 equivalents or less, 2.0 equivalents or less, 1.6 equivalents or less, 1.2 equivalents or less, or 1.1 equivalents or less, relative to the mercapto groups of the polypeptide. By setting the upper limit of the amount of the reducing agent within the above range, the reaction between the (B1) component and the reducing agent and the generation of by-products can be further suppressed while the reaction proceeds.

[0088] In the above production method, the reaction between the polypeptide (component (A)) and the compound (component (B1)) may be carried out, for example, at 55°C or higher. In a situation where the reaction temperature exceeds 55°C, disulfide bond formation between mercapto groups of the polypeptide may usually occur in dimethyl sulfoxide. In contrast, in the production method according to the present disclosure, the reaction is carried out in the presence of a reducing agent, which can inhibit the formation of the disulfide bond. This makes it possible to improve the activity of the reaction between the polypeptide and the compound in a state where the formation of the disulfide bond is suppressed, thereby improving the efficiency of synthesis of the polymer compound.

[0089] In the above production method, the reaction temperature of the reaction between component (A)) and component (B1) may be, for example, 20°C or higher, 30°C or higher, 40°C or higher, 50°C or higher, 53°C or higher, 55°C or higher, 57°C or higher, or 60°C or higher. By setting the lower limit of the reaction temperature within the above range, the reaction between component (A) and component (B1) can be further promoted. In the above production method, the upper limit of the reaction temperature of the reaction between component (A)) and component (B1) may be, for example, 110°C or lower, 100°C or lower, 90°C or lower, 85°C or lower, 80°C or lower, 75°C or lower, or 70°C or lower. By setting the upper limit of the reaction temperature within the above range, the formation of bonds between mercapto groups of the polypeptide as a side reaction can be further suppressed, and the desired polymer compound can be efficiently synthesized. The reaction temperature for the reaction between component (A) and component (B1) may be adjusted within the above-mentioned range, and may be, for example, 50 to 90° C., 53 to 80° C., 55 to 75° C., or 55 to 70° C. The reaction proceeds at any temperature, but a temperature at which the DMSO solvent does not freeze and protein decomposition does not occur is desirable.

[0090] The method for producing a polymer compound may further include other steps, such as separately preparing the component (A) and separately preparing the component (B1).

[0091] The method for preparing component (A) may be a method including expressing a nucleic acid in a host transformed with an expression vector described later. In this case, component (A) may be an artificial fibroin. In addition to direct expression, secretory production, fusion protein expression, etc. may be performed according to the method described in Molecular Cloning, 2nd Edition. When expressed in yeast, animal cells, or insect cells, it can be obtained as a polypeptide to which sugar or a sugar chain is added.

[0092] The artificial fibroin can be produced, for example, by culturing a host transformed with an expression vector in a culture medium, producing and accumulating the artificial fibroin in the culture medium, and collecting it from the culture medium. The method of culturing the host in the culture medium can be carried out according to the method commonly used for culturing the host.

[0093] When the host is a prokaryote such as Escherichia coli or a eukaryote such as yeast, the culture medium for the host may be either a natural medium or a synthetic medium, so long as it contains a carbon source, a nitrogen source, inorganic salts, etc. that can be assimilated by the host and allows efficient cultivation of the host.

[0094] The carbon source may be any that can be assimilated by the host, and examples of the carbon source that can be used include carbohydrates such as glucose, fructose, sucrose, and molasses containing these, starch, and starch hydrolysates, organic acids such as acetic acid and propionic acid, and alcohols such as ethanol and propanol.

[0095] Examples of nitrogen sources that can be used include ammonia, ammonium salts of inorganic or organic acids such as ammonium chloride, ammonium sulfate, ammonium acetate, and ammonium phosphate, other nitrogen-containing compounds, as well as peptone, meat extract, yeast extract, corn steep liquor, casein hydrolysate, soybean meal and soybean meal hydrolysate, various fermentation bacteria and digests thereof.

[0096] Examples of inorganic salts that can be used include monopotassium phosphate, dipotassium phosphate, magnesium phosphate, magnesium sulfate, sodium chloride, ferrous sulfate, manganese sulfate, copper sulfate, and calcium carbonate.

[0097] Cultivation of prokaryotes such as Escherichia coli or eukaryotes such as yeast can be carried out under aerobic conditions, for example, by shaking culture or submerged aeration and stirring culture. The culture temperature is, for example, 15 to 40°C. The culture time is usually 16 hours to 7 days. The pH of the culture medium during culture is preferably maintained at 3.0 to 9.0. The pH of the culture medium can be adjusted using an inorganic acid, an organic acid, an alkaline solution, urea, calcium carbonate, ammonia, or the like.

[0098] During the culture, antibiotics such as ampicillin and tetracycline may be added to the culture medium as necessary. When culturing a microorganism transformed with an expression vector using an inducible promoter as a promoter, an inducer may be added to the medium as necessary. For example, isopropyl-β-D-thiogalactopyranoside or the like may be added to the medium when culturing a microorganism transformed with an expression vector using a lac promoter, and indoleacrylic acid or the like may be added to the medium when culturing a microorganism transformed with an expression vector using a trp promoter.

[0099] Examples of culture media for insect cells that can be used include commonly used TNM-FH medium (Pharmingen), Sf-900 II SFM medium (Life Technologies), ExCell400, ExCell405 (all JRH Biosciences), and Grace's Insect Medium (Nature, 195, 788 (1962)).

[0100] Insect cells can be cultured for 1 to 5 days under conditions such as a culture medium pH of 6 to 7 and a culture temperature of 25 to 30° C. Furthermore, antibiotics such as gentamicin may be added to the culture medium during culture, if necessary.

[0101] When the host is a plant cell, the transformed plant cell may be cultured as it is, or may be differentiated into a plant organ and then cultured. As a medium for culturing the plant cell, for example, commonly used Murashige and Skoog (MS) medium, White medium, or a medium supplemented with plant hormones such as auxin or cytokinin can be used.

[0102] The animal cells can be cultured for 3 to 60 days under conditions such as a culture medium pH of 5 to 9 and a culture temperature of 20 to 40° C. Furthermore, antibiotics such as kanamycin and hygromycin may be added to the medium during culture, if necessary.

[0103] The method of producing artificial fibroin using a host transformed with the above expression vector includes a method of producing the artificial fibroin inside the host cell, a method of secreting the artificial fibroin outside the host cell, and a method of producing the artificial fibroin on the outer membrane of the host cell, etc. Each of these methods can be selected by changing the host cell used and the structure of the artificial fibroin to be produced.

[0104] For example, when artificial fibroin is produced inside a host cell or on the outer membrane of the host cell, the artificial fibroin can be modified to be actively secreted outside the host cell by applying the method of Paulson et al. (J. Biol. Chem., 264, 17619 (1989)), the method of Rowe et al. (Proc. Natl. Acad. Sci. USA, 86, 8227 (1989), Genes Develop., 4, 1288 (1990)), or the methods described in JP-A-05-336963 and WO 1994 / 023021, etc., mutatis mutandis. That is, by using a genetic recombination technique to express a polypeptide containing an active site of the artificial fibroin in a form in which a signal peptide has been added, the artificial fibroin can be actively secreted outside the host cell.

[0105] The artificial fibroin produced by the host transformed with the expression vector can be isolated and purified by the method generally used for isolating and purifying proteins.For example, when the artificial fibroin is expressed in a dissolved state in cells, after the end of the culture, the host cells are collected by centrifugation, suspended in an aqueous buffer solution, and then disrupted by an ultrasonic homogenizer, French press, Manton Gaulin homogenizer, Dyno Mill, etc. to obtain a cell-free extract. The supernatant obtained by centrifuging the cell-free extract can be used to obtain a purified preparation by using, alone or in combination, methods commonly used for isolating and purifying proteins, namely, solvent extraction, salting out with ammonium sulfate or the like, desalting, precipitation with an organic solvent, anion exchange chromatography using resins such as diethylaminoethyl (DEAE)-Sepharose and DIAION HPA-75 (manufactured by Mitsubishi Chemical Corporation), cation exchange chromatography using resins such as S-Sepharose FF (manufactured by Pharmacia), hydrophobic chromatography using resins such as butyl Sepharose and phenyl Sepharose, gel filtration using molecular sieves, affinity chromatography, chromatofocusing, electrophoresis such as isoelectric focusing, and the like.

[0106] As the above-mentioned chromatography, column chromatography using Phenyl-Toyopearl (Tosoh), DEAE-Toyopearl (Tosoh), or Sephadex G-150 (Pharmacia Biotech) is preferably used.

[0107] In addition, when the artificial fibroin is expressed by forming an insoluble body within the cells, the host cells are similarly recovered, then disrupted, and centrifuged to recover the insoluble body of the artificial fibroin as a precipitate fraction. The recovered insoluble body of the artificial fibroin can be solubilized with a protein denaturant. After this operation, a purified sample of the artificial fibroin can be obtained by the same isolation and purification method as above.

[0108] When the artificial fibroin or a derivative in which a sugar chain is added to the artificial fibroin is secreted outside the cells, the artificial fibroin or its derivative can be recovered from the culture supernatant. That is, the culture is treated by a method such as centrifugation to obtain a culture supernatant, and a purified specimen can be obtained from the culture supernatant by the same isolation and purification method as described above.

[0109] The component (A) may be an artificial fibroin. When the artificial fibroin is prepared separately, the method may be, for example, a method of artificially synthesizing naturally derived fibroin. Here, examples of naturally derived fibroin include fibroin produced by insects or spiders. Natural fibroin is a fibrous protein with a molecular weight of about 370,000, composed of two subunits, and has a high content of glycine residues, alanine residues, serine residues, and tyrosine residues, with these amino acid residues accounting for nearly 90% of the total number of amino acid residues. Natural fibroin has a crystalline region rich in amino acid residues with relatively small side chains such as glycine, alanine, and serine, and an amorphous region with amino acid residues with relatively large side chains such as tyrosine.

[0110] More specific examples of naturally derived fibroins include fibroins whose sequence information is registered in NCBI GenBank. For example, from among the sequences registered in NCBI GenBank that contain INV as DIVISION, it can be confirmed by extracting sequences with spidroin, ampullate, fibroin, "silk and polypeptide," or "silk and protein" as keywords in DEFINITION, a specific product character string in CDS, and a specific character string in TISSUE TYPE in SOURCE.

[0111] In this specification, "artificial fibroin" means artificially produced fibroin (artificial fibroin). The artificial fibroin may be a fibroin having an amino acid sequence different from that of naturally occurring fibroin, or may be a fibroin having an amino acid sequence identical to that of naturally occurring fibroin. The artificial fibroin can be produced by a known method, for example, by the method described in International Publication No. 2019 / 194263.

[0112] The artificial fibroin may be a fibrous protein having a structure similar to that of naturally occurring fibroin, or may be a fibroin having a sequence similar to the repetitive sequence of naturally occurring fibroin. The "similar sequence to the repetitive sequence of fibroin" may be a sequence that is actually found in naturally occurring fibroin, or a sequence similar thereto.

[0113] The "artificial fibroin" may be one whose amino acid sequence has been modified based on naturally occurring fibroin (for example, one whose amino acid sequence has been modified by modifying the gene sequence of cloned naturally occurring fibroin) as long as it has the amino acid sequence specified in the present disclosure, or one whose amino acid sequence has been artificially designed without relying on naturally occurring fibroin (for example, one whose desired amino acid sequence has been obtained by chemically synthesizing a nucleic acid encoding a designed amino acid sequence). Note that artificial fibroin whose amino acid sequence has been modified is also included in the artificial fibroin as long as the amino acid sequence is different from that of naturally occurring fibroin. Examples of artificial fibroin include artificial silk fibroin (one whose amino acid sequence of silk protein produced by silkworms has been modified) and artificial spider silk fibroin (one whose amino acid sequence of spider silk protein produced by spiders has been modified). Since artificial fibroin is relatively easy to fibrillate and has a high fiber forming ability, it is preferable for the molding material to include artificial spider silk fibroin, and more preferably to consist of artificial spider silk fibroin.

[0114] The artificial fibroin according to this embodiment may be a protein containing a domain sequence represented by formula 1: [(A)n motif-REP]m, or formula 2: [(A)n motif-REP]m-(A)n motif. The artificial fibroin may further have amino acid sequences (N-terminal sequence and C-terminal sequence) added to either or both of the N-terminal and C-terminal sides of the domain sequence. The N-terminal sequence and C-terminal sequence are typically, but are not limited to, regions that do not have repetitions of amino acid motifs characteristic of fibroin, and consist of about 100 amino acid residues.

[0115] In the present specification, the term "domain sequence" refers to an amino acid sequence represented by formula 1: [(A)n motif-REP]m or formula 2: [(A)n motif-REP]m-(A)n motif. Here, the (A)n motif indicates an amino acid sequence mainly composed of alanine residues, and the number of amino acid residues is 2 to 27. The number of amino acid residues in the (A)n motif may be an integer of 2 to 20, 4 to 27, 4 to 20, 8 to 20, 10 to 20, 4 to 16, 8 to 16, or 10 to 16. In addition, the ratio of the number of alanine residues to the total number of amino acid residues in the (A)n motif may be 40% or more, and may be 60% or more, 70% or more, 80% or more, 83% or more, 85% or more, 86% or more, 90% or more, 95% or more, or 100% (meaning that it is composed only of alanine residues). At least seven of the (A)n motifs present in the domain sequence may be composed only of alanine residues. REP represents an amino acid sequence consisting of 2 to 200 amino acid residues. REP may be an amino acid sequence consisting of 10 to 200 amino acid residues. m represents an integer of 2 to 300, and may be an integer of 10 to 300. A plurality of (A)n motifs may be the same amino acid sequence as each other or different amino acid sequences. A plurality of REPs may be the same amino acid sequence as each other or different amino acid sequences.

[0116] Specific examples of artificial fibroins include artificial fibroins derived from the major ampullate gland of spiders (first artificial fibroin), artificial fibroins having a domain sequence with a reduced content of glycine residues (second artificial fibroin), artificial fibroins having a domain sequence with a reduced content of (A)n motifs (third artificial fibroin), artificial fibroins having a reduced content of glycine residues and a reduced content of (A)n motifs (fourth artificial fibroin), artificial fibroins having a domain sequence including a region with a locally high hydrophobic index (fifth artificial fibroin), and artificial fibroins having a domain sequence with a reduced content of glutamine residues (sixth artificial fibroin). The definitions of the first to sixth artificial fibroins are incorporated herein by reference in the contents of WO 2019 / 194263.

[0117] The artificial fibroin may contain a tag sequence at either or both of the N-terminus and C-terminus, which allows the artificial fibroin to be isolated, immobilized, detected, visualized, and the like.

[0118] An example of a tag sequence is an affinity tag that utilizes specific affinity (binding ability, affinity) with other molecules. A specific example of an affinity tag is a histidine tag (His tag). The His tag is a short peptide consisting of about 4 to 10 histidine residues lined up, and has the property of specifically binding to metal ions such as nickel, and can be used to isolate artificial fibroin by chelating metal chromatography. A specific example of a tag sequence is the amino acid sequence shown in SEQ ID NO: 8 (an amino acid sequence including a His tag sequence and a hinge sequence).

[0119] Furthermore, tag sequences such as glutathione-S-transferase (GST) that specifically binds to glutathione, and maltose-binding protein (MBP) that specifically binds to maltose can also be used.

[0120] Furthermore, an "epitope tag" that utilizes an antigen-antibody reaction can also be used. By adding an antigenic peptide (epitope) as a tag sequence, an antibody against the epitope can be bound. Examples of epitope tags include HA (peptide sequence of influenza virus hemagglutinin) tag, myc tag, and FLAG tag. By using an epitope tag, artificial fibroin can be easily purified with high specificity.

[0121] Furthermore, a tag sequence that can be cleaved with a specific protease can also be used. By treating the protein adsorbed via the tag sequence with the protease, the artificial fibroin from which the tag sequence has been cleaved can be collected.

[0122] Specific examples of the artificial fibroin include the artificial fibroins shown in Table 1.

[0123] [Table 1]

[0124] The artificial fibroin may be an artificial fibroin having at least two or more characteristics of the first artificial fibroin, the second artificial fibroin, the third artificial fibroin, the fourth artificial fibroin, the fifth artificial fibroin, and the sixth artificial fibroin.

[0125] The molecular weight of the artificial fibroin is not particularly limited, but may be, for example, 2 kDa or more and 700 kDa or less. The molecular weight of the artificial fibroin according to the present embodiment may be, for example, 2 kDa or more, 3 kDa or more, 4 kDa or more, 5 kDa or more, 6 kDa or more, 7 kDa or more, 8 kDa or more, 9 kDa or more, 10 kDa or more, 20 kDa or more, 30 kDa or more, 40 kDa or more, 50 kDa or more, 60 kDa or more, 70 kDa or more, 80 kDa or more, 90 kDa or more, or 100 kDa or more, and may be 700 kDa or less, 600 kDa or less, 500 kDa or less, 400 kDa or less, less than 360 kDa, 300 kDa or less, or 200 kDa or less.

[0126] As described above, when obtaining a molded body having excellent flexibility using the synthetic polymer obtained by the above-mentioned production method, if the component (B1) is a low molecular weight compound, it is preferable to use a polypeptide having a relatively low molecular weight as the component (A). However, when the polypeptide has a low molecular weight (for example, 30 kDa or less), the yield may decrease during purification in the preparation. In particular, when a purification method for a hydrophobic protein is used, there is a risk of problems such as a decrease in purity. Therefore, in the above-mentioned production method, for example, a protein (PRT2882) having an amino acid sequence shown in SEQ ID NO: 9 in which a hydrophobic tag (for example, GFILGFIL in SEQ ID NO: 9) is introduced at the N-terminus or C-terminus may be used. Since a low molecular weight recombinant protein having an amino acid sequence including such a hydrophobic tag sequence can be obtained in high yield and purity during purification, the above-mentioned production method can obtain a polymer compound having a low content of impurities. As a result, in a molded body using such a polymer compound as a molding material, merits such as better mechanical properties can be advantageously secured.

[0127] In the artificial fibroin according to this embodiment, one or more of the amino acid residues constituting the REP may be hydrophobic amino acid residues. In other words, it is preferable that the REP contains a hydrophobic amino acid residue. The hydrophobic amino acid residue means an amino acid residue with a positive hydrophobic index. The hydrophobic index (Hydropathy index, hereinafter also referred to as "HI") of the amino acid residue is a known index (Hydropathy index: Kyte J, & Doolittle R (1982) "A simple method for displaying the hydropathic character of a protein", J. Mol. Biol., 157, pp. 105-132). Examples of hydrophobic amino acid residues include isoleucine (HI: 4.5), valine (HI: 4.2), leucine (HI: 3.8), phenylalanine (HI: 2.8), methionine (HI: 1.9), and alanine (HI: 1.8).

[0128] In the artificial fibroin according to this embodiment, it is preferable that the domain sequence has an amino acid sequence corresponding to the insertion of a cysteine ​​residue in REP, as compared with naturally occurring fibroin.

[0129] The domain sequence preferably has an amino acid sequence corresponding to a cysteine ​​residue inserted adjacent to a glycine, serine or alanine residue in the REP, more preferably an amino acid sequence corresponding to a cysteine ​​residue inserted adjacent to a glycine residue in the REP. The cysteine ​​residue in the REP may be located between the glycine, serine or alanine residue and the glycine, serine or alanine residue, or between the serine and glycine residue.

[0130] The domain sequence preferably has an amino acid sequence corresponding to the insertion of a cysteine ​​residue at a position adjacent to the hydrophobic amino acid residue in the REP. In this case, the hydrophobic amino acid residues are fixed between the molecules by hydrophobic interaction. The cysteine ​​residue in the REP may be located next to the hydrophobic amino acid residue, may be located between the hydrophobic amino acid residue and an amino acid residue other than the hydrophobic amino acid residue, may be located between the hydrophobic amino acid residue and a glycine residue, a serine residue, or an alanine residue, or may be located between the hydrophobic amino acid residue and a glycine residue. The hydrophobic amino acid residue may be one selected from the group consisting of an isoleucine residue, a valine residue, a leucine residue, a phenylalanine residue, a methionine residue, and an alanine residue.

[0131] The domain sequence may have an amino acid sequence corresponding to the insertion of a cysteine ​​residue in the REP located near the N-terminus and / or the C-terminus of the domain sequence, as compared with naturally occurring fibroin. In this case, the molecular chain can be lengthened. In this specification, the REP located near the N-terminus of the domain sequence means the REP located at the 1st to 3rd positions from the N-terminus of the domain sequence. For example, the cysteine ​​residue may be located in the REP located at the 1st to 2nd positions from the N-terminus of the domain sequence. In this specification, the REP located near the C-terminus of the domain sequence means the REP located at the 1st to 3rd positions from the C-terminus of the domain sequence. For example, the cysteine ​​residue may be located in the REP located at the 1st to 2nd positions from the C-terminus of the domain sequence. It is preferable that the cysteine ​​residue is located in the REP located at the most N-terminus side and / or the most C-terminus side of the domain sequence.

[0132] The domain sequence may have an amino acid sequence corresponding to the insertion of a cysteine ​​residue in the center or near the center in the REP, compared with naturally occurring fibroin. In this specification, near the center of the amino acid sequence in the REP refers to the 1st to 5th positions from the amino acid residue located in the center of the REP (the amino acid residue on the N-terminal side when there are two amino acid residues located in the center) toward the N-terminal side, or the 1st to 5th positions from the amino acid residue located in the center of the REP (the amino acid residue on the C-terminal side when there are two amino acid residues located in the center). For example, the cysteine ​​residue may be located in the center of the REP, or may be located in the 1st to 3rd or 1st to 2nd positions from the amino acid residue located in the center of the REP toward the N-terminal side or C-terminal side. Here, when a first segment is used that includes a polypeptide backbone in which cysteine ​​residues are inserted so that, for example, one cysteine ​​residue is located at each of the N-terminal side and the C-terminal side of the domain sequence, a synthetic polymer can be obtained in which such first segments and second segments are linked so that one cysteine ​​residue is located alternately. A molded product (for example, a fiber, a film, a gel, etc.) obtained using this synthetic polymer can be expected to have improved elongation. Furthermore, when a first segment is used that includes a polypeptide backbone in which a cysteine ​​residue is inserted, for example, so that the cysteine ​​residue is located closer to the center of the domain sequence than the N-terminus or C-terminus, a synthetic molecule in which a second segment is linked to such a first segment can be expected to have improved solubility in a solvent.

[0133] The hydrophobicity (hydropathy index) of the REP of the artificial fibroin may be, for example, -0.80, -0.70, -0.06 or more, -0.50 or more, -0.40 or more, -0.30 or more, -0.20 or more, -0.10 or more, 0.00 or more, 0.10 or more, 0.20 or more, 0.22 or more, 0.25 or more, 0.30 or more, 0.35 or more, 0.40 or more, 0.45 or more, 0.50 or more, 0.55 or more, 0.60 or more, 0.65 or more, or 0.70 or more. There is no particular upper limit to the hydrophobicity of the REP, and it may be 1.0 or less, or 0.7 or less.

[0134] In the present specification, the "hydrophobicity of REP" is a value calculated by the following method. Formula 1: [(A) n Motif-REP] m Or formula 2: [(A) n Motif-REP] m -(A) n In the fibroin containing the domain sequence represented by the motif, it is located at the most C-terminal side (A). n For all REPs contained in the sequence excluding the sequence from the motif to the C-terminus of the domain sequence from the domain sequence, the sum of the hydrophobicity indexes of each amino acid residue in that region is e, and the hydrophobicity index located at the most C-terminus side (A) is n The sequence from the motif to the C-terminus of the domain sequence is removed from the domain sequence, and further (A) n When the total number of amino acid residues of all REPs excluding the motif is f, the hydrophobicity of the REP is calculated as e / f. n The reason for targeting "the sequence obtained by excluding the sequence from the motif to the C-terminus of the domain sequence from the domain sequence" is the same as that mentioned above.

[0135] The domain sequence may have an amino acid sequence corresponding to 1 to less than 16 cysteine ​​residues inserted into REP compared to naturally-occurring fibroin. In other words, the total number of cysteine ​​residues corresponding to insertion into REP may be 1 to less than 16. The total number of cysteine ​​residues corresponding to insertion into REP may be 1 to 12, 1 to 10, 1 to 8, 1 to 6, or 2 to 4. The number of cysteine ​​residues per REP in the domain sequence may be, for example, 1 to 3, 1 to 2, or 1.

[0136] The total number of cysteine ​​residues in the artificial fibroin of this embodiment may be 1 or more and less than 16, 1 or more and 12 or less, 1 or more and 10 or less, 1 or more and 8 or less, 1 or more and 6 or less, or 2 or more and 4 or less.

[0137] In addition to the modifications of the cysteine ​​residues in the REP described above, the artificial fibroin of this embodiment may further have modifications in the amino acid sequence equivalent to the substitution, deletion, insertion and / or addition of one or more amino acid residues compared to naturally occurring fibroin.

[0138] The artificial fibroin according to this embodiment preferably has an amino acid sequence corresponding to the insertion of a cysteine ​​residue.

[0139] The method for preparing the component (B1) may be a method including esterifying maleic anhydride with at least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates. In this case, an acid or a base may be used, and the reaction may be carried out without a solvent. In the preparation of the component (B1), it is not necessary to introduce a protective group into the polyether, and a simple method can be adopted. The purification may be carried out by washing with a solvent such as cyclopentyl methyl ether, tetrahydrofuran, ethyl acetate, or ethanol to remove impurities.

[0140] The second embodiment of the method for producing a polymer compound comprises reacting a polypeptide having at least one mercapto group with at least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, which has two structures represented by the following general formula (2) (hereinafter also referred to as component (B2)), in dimethyl sulfoxide in the presence of a base, a reducing agent, and a polymerization inhibitor (hereinafter also referred to as component (E)). 1represents a hydrogen atom or a methyl group. The heating reaction is a reaction between component (A) and component (B2), and is carried out, for example, by heating to 50° C. or higher. The timing of heating is not limited to after components (A) to (E) are mixed. For example, components (A) to (E) may be added to preheated dimethyl sulfoxide, components (C), (D), and (E) may be added to dimethyl sulfoxide, heating may be started, and after a predetermined temperature is reached, components (A) and (B) may be added and reacted, or one of components (A) and (B) and components (C), (D), and (E) may be added, and after a predetermined temperature is reached, the remaining one of components (A) and (B) may be added and reacted.

[0141] [ka]

[0142] In the following description, the differences from the first embodiment will be described, and the description of the polypeptide having at least one mercapto group, the base, and the reducing agent in the second embodiment can be applied to the components (A), (C), and (D) in the first embodiment.

[0143] At least one compound (component (B2)) selected from the group consisting of polyethers, polyesters, and polycarbonates, which has two structures represented by general formula (2), has a structure represented by formula (1) and is therefore capable of reacting with a mercapto group in the above-mentioned polypeptide, and is a compound capable of introducing a polyether structure, a polyester structure, or a polycarbonate structure into the polypeptide.

[0144] Examples of polyethers having two structures represented by formula (2) include polyethylene glycol, polytetramethylene glycol, polypropylene glycol, and ethylene glycol-propylene glycol copolymers. Examples of polyesters having two structures represented by formula (2) include polylactic acid, polyglycolic acid, polybutylene succinate, polycaprolactone, and polyhydroxyalkanoates including 3-hydroxybutanoic acid-3-hydroxyhexanoic acid copolymers. Examples of polycarbonates having two structures represented by formula (2) include aliphatic polycarbonates such as polyethylene carbonate, polypropylene carbonate, and polytrimethylene carbonate. These compounds have easier molecular motion than the above-mentioned polypeptides, and therefore, when they are the above-mentioned compounds, the flexibility of the molded article made of the obtained polymer compound can be further improved.

[0145] Regarding the amount of the component (B2) to be used, the explanation given for the component (B1) in the first embodiment above can be read as being for the component (B2) and applied.

[0146] The polymerization inhibitor (component (E)) in the above-mentioned production method is a compound that inhibits the self-polymerization of component (B2). Examples of component (E) include hydroquinone, methoquinone, 4-tert-butylpyrocatechol, tert-butylhydroquinone, 1,4-benzoquinone, dibutylhydroxytoluene, methoquinol, phenothiazine, and 1,1-diphenyl-2-picrylhydrazyl.

[0147] The content of the (E) component may be, for example, 0.01 parts by mass or more, 0.05 parts by mass or more, 0.1 parts by mass or more, or 0.2 parts by mass or more, relative to 100 parts by mass of the above compound (component (B2)). By setting the content of the polymerization inhibitor within the above range, the initiation of radical polymerization of the structure represented by general formula (2) is suppressed, and it may be possible to make the reaction between the structure represented by general formula (2) and the mercapto group of the polypeptide more dominant. The content of the (E) component may be, for example, 25 parts by mass or less, 10 parts by mass or less, 5 parts by mass or less, or 1 part by mass or less, relative to 100 parts by mass of the (B2) component. By setting the content of the (E) component within the above range, it is possible to further suppress the (E) component from inhibiting the reaction in the system.

[0148] The method for preparing the component (B2) may be a method including esterifying (meth)acrylic acid, (meth)acrylic anhydride, (meth)acryloyl chloride, and at least one compound selected from the group consisting of polyether, polyester, and polycarbonate. The preparation of the component (B2) does not require the introduction of a protective group into the polyether, and a simple method can be adopted. The purification was performed by removing organic salts by hot extraction with tetrahydrofuran, and further adding n-hexane to cause reprecipitation.

[0149] The polymer compound synthesized by the above-mentioned production method has a structure in which a polypeptide portion and at least one structural unit selected from the group consisting of a polyether structure, a polyester structure, and a polycarbonate structure are directly bonded to each other through a structure represented by the following general formula (3), (4), or (5) such that the polypeptide portion is bonded to S in the following general formula (3), (4), or (5). In general formula (3) and general formula (4), M represents any one of H (hydrogen atom), Na (sodium atom), K (potassium atom), NHEt3, or NHEtiPr2, and in general formula (5), R 1 represents H or Me. The above Me represents a methyl group, the above Et represents an ethyl group, and the above iPr represents an isopropyl group.

[0150] [ka]

[0151] [ka]

[0152] [ka]

[0153] The above-mentioned polymer compound can also be called a block copolymer since it has a polypeptide and at least one structural unit selected from the group consisting of a polyether structure, a polyester structure, and a polycarbonate structure. When a structural unit consisting of a polypeptide is A and a structural unit containing at least one of a polyether structure, a polyester structure, and a polycarbonate structure is B, the above-mentioned polymer compound only needs to have at least one each of A and B. The above-mentioned polymer compound may be, for example, an AB diblock copolymer, an ABA triblock copolymer, an ABAB tetrablock copolymer, an ABABA pentablock copolymer, or a copolymer such as AABBBABBAA.

[0154] The polymer compound may be a linear polymer or a comb polymer, or may have a three-dimensional network structure. From the viewpoint of further improving the flexibility of a molded article made of the polymer compound, the polymer compound is preferably a linear polymer or a comb polymer, and from the viewpoint of improving the elastic modulus of the molded article, the polymer compound is preferably a three-dimensional network structure.

[0155] The position of at least one structural unit selected from the group consisting of a polyether structure, a polyester structure, and a polycarbonate structure to be introduced into the polypeptide portion is not particularly limited, and may be the end of the polypeptide portion or a position other than the end of the polypeptide portion. However, from the perspective of ease of production of the polymer compound, the end of the polypeptide portion is preferable.

[0156] The polymer compound synthesized by the above-mentioned production method can be used as a molding material. The shape of the molded product using the above-mentioned polymer compound as a molding material is not particularly limited, and may be, for example, a film, fiber, etc.

[0157] One aspect of the present disclosure provides a method for producing a polymer solution, which includes dissolving the polymer obtained by the above-mentioned production method in a solvent. The polymer solution obtained by such a method can be used as a so-called dope solution when preparing a molded body. Examples of solvents used for preparing the dope solution include formic acid, dimethyl sulfoxide, N,N-dimethylformamide, N,N-dimethylacetamide, hexafluoro-2-propanol, and N-methyl-2-pyrrolidone. In the above-mentioned production method, the target polymer is obtained in a state dissolved in dimethyl sulfoxide (as a dimethyl sulfoxide solution) by the above-mentioned heating reaction. Therefore, the above-mentioned production method for the polymer solution includes a step of dissolving the polymer extracted from dimethyl sulfoxide by, for example, a known extraction method or separation method in a predetermined solvent.

[0158] A polymer film may be produced by a method including a step of forming a film from the polymer solution obtained by the above-mentioned method for producing a polymer solution. The forming into the film may be, for example, cast molding.

[0159] Polymer fibers may be produced by a method including a step of spinning the polymer solution obtained by the above-mentioned method for producing a polymer solution.

[0160] The dimethylsulfoxide solution of the polymer compound obtained by the heat reaction in the above-mentioned method for producing the polymer compound can be used as it is as a dope for producing a film or fiber. The polymer film can be produced more simply and at low cost. The film can be formed by, for example, cast molding.

[0161] One aspect of the present disclosure provides a method for producing a film, comprising: reacting a polypeptide having at least one mercapto group with at least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, having two structures represented by the following general formula (1), in dimethyl sulfoxide in the presence of a base and a reducing agent to obtain a polymer solution (a dimethyl sulfoxide solution of a polymer), and forming a film from the polymer solution. In general formula (1), M represents any one of H, Na, K, NHEt3, and NHEtiPr2.

[0162] [ka]

[0163] One aspect of the present disclosure provides a method for producing a film, comprising: reacting a polypeptide having at least one mercapto group with at least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, which has two structures represented by the following general formula (2), in dimethyl sulfoxide in the presence of a base, a reducing agent, and a polymerization inhibitor to obtain a polymer compound solution (a dimethyl sulfoxide solution of a polymer compound), and forming a film from the polymer compound solution. 1 represents a hydrogen atom or a methyl group.

[0164] [ka]

[0165] One aspect of the present disclosure provides a method for producing a fiber, comprising: reacting a polypeptide having at least one mercapto group with at least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, the compound having two structures represented by the following general formula (1), in dimethyl sulfoxide in the presence of a base and a reducing agent, to obtain a polymer solution (a dimethyl sulfoxide solution of a polymer), and spinning the polymer solution. In general formula (1), M represents any of H, Na, K, NHEt3, and NHEtiPr2.

[0166] [ka]

[0167] One aspect of the present disclosure provides a method for producing fibers, comprising: reacting a polypeptide having at least one mercapto group with at least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, the compound having two structures represented by the following general formula (2), in dimethyl sulfoxide in the presence of a base, a reducing agent, and a polymerization inhibitor, to obtain a polymer solution (a dimethyl sulfoxide solution of a polymer), and spinning the polymer solution. 1 represents a hydrogen atom or a methyl group.

[0168] [ka]

[0169] One aspect of the present disclosure provides a film comprising a polymer compound, the polymer compound having a structure in which a polypeptide portion and at least one structural unit selected from the group consisting of a polyether structure, a polyester structure, and a polycarbonate structure are directly bonded to each other via a structure represented by the following formula (3), (4), or (5) such that the polypeptide portion is bonded to S in the following formula (3), (4), or general formula (5), the film having an elongation of 400% or more and a ratio of breaking strength to yield strength of more than 1. In general formulas (3) and (4), M represents any one of H, Na, K, NHEt3, and NHEtiPr2, and in general formula (5), R 1 represents H or Me.

[0170] [ka]

[0171] [ka]

[0172] [ka]

[0173] One aspect of the present disclosure provides a fiber comprising a polymer compound, the polymer compound having a structure in which a polypeptide portion and at least one structural unit selected from the group consisting of a polyether structure, a polyester structure, and a polycarbonate structure are directly bonded via a structure represented by the following formula (3), (4), or (5) such that the polypeptide portion is bonded to S in the following formula (3), (4), or general formula (5), the fiber having an elongation of 400% or more and a ratio of breaking strength to yield strength of more than 1. In general formula (5), R1 represents a hydrogen atom or a methyl group.

[0174] [ka]

[0175] [ka]

[0176] [ka]

[0177] In the above film and fiber, the ratio of breaking strength to yield strength (breaking strength [MPa] / yield strength [MPa]) is greater than 1, and may be, for example, 1.1 or more, 1.2 or more, 1.3 or more, 1.4 or more, or 1.5 or more. In the above film and fiber, the ratio of breaking strength to yield strength (breaking strength [MPa] / yield strength [MPa]) may be, for example, 3 or less, or 2 or less.

[0178] The polymer compound according to the present embodiment can be used as an adhesive for adhering an adherend or as a coating agent for laminating and forming a coating layer on a substrate, in the form of a solution, aqueous dispersion, film, or powder, containing the polymer compound as a main component. These solution-like adhesives and coating agents, water-dispersible adhesives and coating agents, film-like adhesives and coating agents, and powder-like adhesives and coating agents can be obtained, for example, by the following manufacturing methods.

[0179] A solution-type adhesive or solution-type coating agent containing a polymer compound can be manufactured by a method including a step of dissolving the polymer compound obtained by the method of this embodiment in a solvent. According to such a method, unlike conventional solution-type adhesives or solution-type coating agents containing petroleum-derived components, a solution-type adhesive or solution-type coating agent having biodegradability can be easily obtained. In the manufacturing method of the polymer compound according to the present embodiment described above, the target polymer compound is obtained in a state dissolved in dimethyl sulfoxide (as a dimethyl sulfoxide solution) by the above-mentioned heating reaction. Therefore, the manufacturing method of the solution-type adhesive or coating agent according to this embodiment includes a step of dissolving the polymer compound extracted from dimethyl sulfoxide by, for example, a known extraction method or separation method in a predetermined solvent. In addition, the solution-type adhesive or solution-type coating agent according to this embodiment may be composed of a dimethyl sulfoxide solution of the polymer compound obtained by the heating reaction in the manufacturing method of the polymer compound.

[0180] Solvents used in the production of solution adhesives and solution coating agents include aqueous media such as water in which polymer compounds can be dissolved, basic aqueous solutions, acidic aqueous solutions, and neutral aqueous solutions containing inorganic salts, as well as organic solvents, etc. Examples of organic solvents include formic acid, dimethyl sulfoxide (DMSO), hexafluoroisopropanol (HFIP), N,N-dimethylformamide (DMF), N,N-dimethylacetamide (DMAc), N-methylpiperidone (NMP), and dihydrolevoglucosenone.

[0181] The polymer compound contained in the solution adhesive or solution coating agent is obtained by the manufacturing method according to this embodiment, and is not particularly limited as long as it is soluble in the above-mentioned aqueous medium or organic solvent. The adhesive or coating agent in which the polymer compound is dissolved in an aqueous medium has the advantage of being easier to handle than the adhesive or coating agent in which the polymer compound is dissolved in an organic solvent. The polymer compound according to this embodiment has a molecule that can plasticize the protein bound to the protein, and therefore exhibits higher water solubility than the protein. Therefore, the aqueous solution adhesive or aqueous solution coating agent in which the polymer compound is dissolved in an aqueous medium can have a higher protein content than the aqueous solution adhesive or aqueous solution coating agent in which the protein is simply dissolved in an aqueous medium. Therefore, when an aqueous solution adhesive or aqueous solution coating agent in which a polymer compound is dissolved in an aqueous medium is used, the adhesive strength of the adhesive or coating agent to the adherend surface of the adherend or the laminate surface described below can be expected to be improved compared to the case in which an aqueous solution adhesive or aqueous solution coating agent in which a protein is dissolved in an aqueous medium is used.

[0182] The concentration of the polymer compound in the solution adhesive or solution coating agent is appropriately determined based on the solubility of the polymer compound in the solvent, etc. Such a concentration may be, for example, 5 to 40 mass%, 10 to 35 mass%, 15 to 20 mass%, or 20 to 25 mass%. In addition, the solution adhesive or solution coating agent may contain components other than the polymer compound, such as various additives contained in known solution adhesives or solution coating agents, as necessary. The upper limit of the concentration may also be, for example, less than 30 mass%, 28 mass% or less, 25 mass% or less, or 23 mass% or less.

[0183] A solution-type adhesive containing a polymer compound is used to produce an adhesive body in which a plurality of adherends are bonded to each other. For example, after the solution-type adhesive is interposed between a plurality of adherends to be bonded, the solvent in the solution-type adhesive is removed and the polymer compound is solidified, thereby bonding the adherends to each other to obtain an adhesive body. According to such a method, an adhesive body can be obtained using a biodegradable adhesive, and there is an advantage that an adhesive body that can reduce the environmental load at the time of disposal can be easily produced. Note that the material of the adherend is not particularly limited as long as it can be bonded with a solution-type adhesive containing a polymer compound, and it may be an organic substance (for example, cellulose products such as paper and wood, synthetic resins, and protein products) or an inorganic substance (non-metallic substances such as metal and glass).

[0184] The specific method for producing an adhesive body using a solution-type adhesive is not limited in any way. For example, the solution-type adhesive may be applied or dropped onto at least one of the adherends to be bonded to each other, or the adherend surface of the adherend may be brought into contact with or immersed in the liquid surface of the solution-type adhesive to make the solution-type adhesive present on the adherend surface, and then the adherends may be overlapped or butted together with their adherend surfaces to intervene between the adherends. In addition, when removing the solvent in the solution-type adhesive intervening between the adherends to solidify the polymer compound, for example, the adherends may be heated, air-dried, or naturally dried while applying pressure from at least one of the overlapping or butting directions of the adherend surfaces so that the overlapped or butted adherend surfaces are in close contact with each other. Of course, if adhesion is possible without applying such pressure, there is no need to apply pressure.

[0185] The solution-type coating agent containing a polymer compound is used to manufacture a laminate in which a coating layer is formed on the entire or part of the surface of a substrate. For example, the solution-type coating agent is supplied to at least a part of the surface of the substrate on which the coating layer is to be laminated, and the surface part of the substrate is coated with the solution-type coating agent, and then the solvent in the solution-type adhesive is removed to solidify the polymer compound. In this way, a laminate can be obtained by forming a coating layer on at least a part of the surface of the substrate. This method also has the advantage that a laminate that can reduce the environmental load at the time of disposal can be easily manufactured because a laminate can be obtained using a coating agent having biodegradability. In addition, the material of the substrate is not particularly limited as long as it is capable of adhering to the solution-type coating agent containing a polymer compound, and may be the same as the adherend to which the solution-type adhesive is adhered.

[0186] The specific method for producing a laminate using a solution-type coating agent is not limited at all. For example, the solution-type coating agent may be applied or dropped onto at least a part of the substrate surface, or at least a part of the substrate surface may be brought into contact with or immersed in the liquid surface of the solution-type coating agent, so that the solution-type coating agent is supplied to at least a part of the substrate to form a layer of a predetermined thickness. In addition, when solidifying the polymer compound in the coating agent layer (solution-type coating agent) laminated on the substrate surface, the coating agent layer on the substrate surface may be heated, air-dried, or naturally dried to remove the solvent in the coating agent layer. In this case, if necessary, a predetermined pressing body (pressurizing body) may be placed on the coating agent layer so as to cover the entire surface of the coating agent layer, and the coating agent layer may be pressed (pressurized) toward the substrate surface so that the coating agent layer is in close contact with the substrate surface. At this time, it is preferable that the pressing body does not adhere to the solution-type coating agent. For example, adhesion of the solution-type coating agent to the pressing body can be prevented by applying a release agent to the contact surface of the pressing body with the coating agent layer, attaching release paper to the contact surface, or performing a surface treatment on the contact surface so that the coating agent will not adhere to the contact surface, or using a pressing body made of a material that will not adhere to the solution-type coating agent.

[0187] A water-dispersible adhesive or water-dispersible coating agent containing a polymer compound can be produced by a method including a step of dispersing the polymer compound obtained by the method of this embodiment in an aqueous medium. According to such a method, unlike conventional water-dispersible adhesives or water-dispersible coating agents containing petroleum-derived components, a water-dispersible adhesive or water-dispersible coating agent having biodegradability can be easily obtained. As described above, in the method for producing a polymer compound according to this embodiment, the target polymer compound is obtained in a state dissolved in dimethyl sulfoxide (as a dimethyl sulfoxide solution). Therefore, in the method for producing a water-dispersible adhesive or water-dispersible coating agent according to this embodiment, a step of dispersing a polymer compound extracted from dimethyl sulfoxide by, for example, a known extraction method or separation method, in a predetermined aqueous medium is included.

[0188] Examples of the aqueous medium used in the production of the water-dispersible adhesive or water-dispersible coating agent include water in which a polymer compound can be dispersed, a basic aqueous solution, an acidic aqueous solution, and an aqueous solution containing an inorganic salt.

[0189] The polymer compound contained in the water-dispersible adhesive or water-dispersible coating agent is obtained by the manufacturing method according to this embodiment, and is not particularly limited as long as it is dispersible in the above-mentioned aqueous medium. The content of the polymer compound in the water-dispersible adhesive or water-dispersible coating agent is appropriately determined in consideration of the adhesion of the water-dispersible adhesive to the adherend, and the adhesion or fixation of the water-dispersible coating agent to the substrate. In addition, the water-dispersible adhesive or water-dispersible coating agent may contain components other than the polymer compound, such as various additives contained in known water-dispersible adhesives or water-dispersible coating agents, as necessary.

[0190] In addition, the polymer compound according to the present embodiment has a higher affinity for aqueous media containing water than proteins, since a molecule capable of plasticizing proteins is bound to the protein. Therefore, a water-dispersible adhesive or water-dispersible coating agent containing such a polymer compound can increase the protein content (dispersion amount) and disperse the polymer compound uniformly in the aqueous medium, compared to a water-dispersible adhesive or water-dispersible coating agent simply obtained by dispersing a protein in an aqueous medium. Therefore, when a water-dispersible adhesive or water-dispersible coating agent in which a polymer compound is dispersed in an aqueous medium is used, the adhesive strength of the adhesive or coating agent to the adherend surface of the adherend or the laminate surface described below can be expected to be improved, compared to the case in which a water-dispersible adhesive or water-dispersible coating agent in which a protein is dispersed in an aqueous medium is used.

[0191] The water-dispersible adhesive containing the polymer compound is used to produce an adhesive body in which a plurality of adherends are bonded to each other. For example, after the water-dispersible adhesive is interposed between a plurality of adherends to be bonded, the aqueous medium in the water-dispersible adhesive is removed and the polymer compound is solidified, so that the adherends are bonded to each other to obtain an adhesive body. According to this method, an adhesive body is obtained using a biodegradable adhesive, and there is an advantage that an adhesive body that can reduce the environmental load at the time of disposal can be easily produced. The material of the adherend is not particularly limited as long as it can be bonded with the water-dispersible adhesive containing the polymer compound, and it may be the same as the adherend bonded with the solution-based adhesive containing the polymer compound.

[0192] The specific method for producing an adhesive using a water-dispersible adhesive is not limited in any way. For example, when a water-dispersible adhesive is interposed between a plurality of adherends to be bonded, the same method as that for interposing a water-dispersible adhesive between a plurality of adherends described above may be adopted. In addition, when removing the aqueous medium in the water-dispersible adhesive interposed between the adherends to solidify the polymer compound, the same method as that for removing the solvent in the solution adhesive interposed between the adherends to solidify the polymer compound may be adopted.

[0193] The water-dispersible coating agent containing a polymer compound is used to manufacture a laminate in which a coating layer is formed on the entire or part of the surface of a substrate. For example, the water-dispersible coating agent is supplied to at least a part of the surface of the substrate on which the coating layer is to be laminated, and the surface part of the substrate is coated with the solution-like coating agent, and then the aqueous medium in the water-dispersible adhesive is removed to solidify the polymer compound. This allows a coating layer to be formed on at least a part of the surface of the substrate to obtain a laminate. This method also has the advantage that a laminate that can reduce the environmental load at the time of disposal can be easily manufactured because a biodegradable coating agent is used to obtain a laminate. The material of the substrate is not particularly limited as long as it is capable of adhering to the water-dispersible coating agent containing a polymer compound, and may be the same as the substrate to which the coating layer is formed with the solution-like coating agent described above.

[0194] The specific method for producing a laminate using a water-dispersible coating agent is not limited at all.For example, when the water-dispersible coating agent is supplied to at least a part of the substrate surface, the same method as that for supplying the above-mentioned solution-type coating agent to the substrate surface can be adopted.In addition, when removing the aqueous medium in the water-dispersible coating agent supplied to the substrate surface to solidify the polymer compound, the same method as that for removing the solvent in the solution-type coating agent supplied to the substrate surface to solidify the polymer compound can be adopted.

[0195] A film-like adhesive or film-like coating agent containing a polymer compound can be produced by a method including a step of forming a film from a polymer compound solution in which the polymer compound obtained by the method of this embodiment is dissolved. According to such a method, unlike conventional film-like adhesives or film-like coating agents containing petroleum-derived components, a film-like adhesive or film-like coating agent having biodegradability can be easily obtained. The method for forming the above-mentioned film may be a known cast molding method.

[0196] As a film-forming solution used in the production of a film-like adhesive or a film-like coating agent, for example, a polymer compound solution used as a solution-like adhesive or a solution-like coating agent is preferably used. Therefore, such a film-forming solution may be a dimethyl sulfoxide solution of a polymer compound obtained by the method for producing a polymer compound according to the present embodiment, or may be a solution obtained by dissolving a polymer compound extracted from such dimethyl sulfoxide by, for example, a known extraction method or separation method in a predetermined solvent. When casting a polymer compound solution, known methods and known conditions similar to those used when casting a protein solution may be used.

[0197] The film-like adhesive containing a polymer compound is used to produce an adhesive body in which a plurality of adherends are bonded to each other. For example, the film-like adhesive is swelled or heated to soften it while being interposed between a plurality of adherends to be bonded, and then the film-like adhesive is pressed against the adherends and cured to bond the adherends to each other to obtain an adhesive body. Alternatively, the film-like adhesive may be softened by swelling or heating in advance, interposed between a plurality of adherends, and then the film-like adhesive may be pressed against the adherends and cured. According to these methods, an adhesive body can be obtained using a biodegradable adhesive, and therefore an adhesive body that can reduce the environmental load at the time of disposal can be easily produced. The adherend may be any object that can be bonded with a film-like adhesive containing a polymer compound, and may be the same as an adherend that is bonded with a solution-based adhesive containing a polymer compound.

[0198] The film-like adhesive softens when it absorbs moisture or is heated, and hardens when it is dried or cooled. In addition, when the polymer compound contained in the film-like adhesive has a property that it shrinks when it is contacted with water or heated, especially when it is formed into a molded body, the film-like adhesive also shrinks when it contacts water or is heated. For this reason, when the film-like adhesive containing the polymer compound is swollen or heated and softened and interposed between the adherends, it partially bites into the gaps present on the adherend surface, and especially when the decorative protein has the above-mentioned shrinkage property, it shrinks while biting into the gaps on the adherend surface. When the film-like adhesive hardens in this state, the film-like adhesive and the adherend are bonded to each other due to the anchor effect. In addition, the film-like adhesive exhibits sufficient flexibility by containing as a main component a polymer compound in which a molecule that plasticizes a protein is bonded to the protein. Therefore, such a film-like adhesive exhibits higher flexibility when softened, and therefore it is expected that it will be able to more fully bite into the gaps on the adherend surface, and as a result, even higher adhesive strength can be obtained. Moreover, since the film-like adhesive has excellent flexibility, it can be peeled off from the adherend after adhering to the adherend, and furthermore, it exhibits the excellent function of being able to be reused as a film-like adhesive. When the film-like adhesive is to be reused, it may be washed with water after being peeled off from the adherend. It is also considered that the adhesion of the film-like adhesive to the adherend is due to the formation of hydrogen bonds between the adherend and the polymer compound.

[0199] When the film-like adhesive is swelled and softened, an aqueous liquid such as water or a solution or dispersion containing water, which is absorbed by the film-like adhesive and can shrink the film-like adhesive, is preferably used. For example, the above-mentioned aqueous liquid may be dropped onto the film-like adhesive or a plurality of adherends may be immersed in the aqueous liquid to cause the film-like adhesive to absorb water, thereby swelling and softening the film-like adhesive. In addition, when the film-like adhesive softened by swelling is hardened, for example, a method of heating and drying, air drying, or natural drying may be adopted in a state where the film-like adhesive is interposed between the adherends.

[0200] When the film-like adhesive is heated to soften it, it may be heated before being interposed between the adherends, or the adherends and the film-like adhesive may be heated as a whole after the film-like adhesive is interposed between the adherends. The heating temperature is not particularly limited as long as it is a temperature that can soften the film-like adhesive and does not adversely affect (e.g., does not decompose) the protein contained in the film-like adhesive. When hardening the film-like adhesive softened by heating, a method of cooling with a cooling device or leaving it to cool can be used.

[0201] In order to allow the film-like adhesive softened by swelling or heating to fully penetrate into the gaps in the adherend's surface, it is desirable to bring the interface between the softened film-like adhesive and the adherend into intimate contact. Therefore, for example, it is preferable to apply pressure to multiple adherends from at least one side in the overlapping or butting direction of the adherend's surfaces, and to press the softened film-like adhesive against the adherend while hardening it. Any common method can be used as the method for applying pressure to the adherend.

[0202] The film-like coating agent containing a polymer compound is used to manufacture a laminate in which a coating layer is formed on the entire or part of the surface of a substrate. For example, the film-like coating agent is placed so as to cover at least a part of the surface of the substrate, and the film-like coating agent is swelled or heated to soften it, and then the film-like coating agent is pressed against the substrate surface and hardened to obtain a laminate by forming a coating layer on at least a part of the substrate surface. Alternatively, the film-like coating agent may be softened by swelling or heating in advance, and then placed so as to cover at least a part of the surface of the substrate, and then hardened in a state in which the film-like coating agent is pressed against the adherend. According to these methods, since a laminate is obtained using a coating agent having biodegradability, there is an advantage that a laminate capable of reducing the environmental load at the time of disposal can be easily manufactured. The substrate may be one to which the film-like coating agent containing a polymer compound can be adhered, and may be the same as the adherend to which the film-like adhesive containing a polymer compound is adhered.

[0203] The mechanism by which the film-like coating agent adheres to the substrate surface is considered to be similar to the mechanism by which the film-like adhesive adheres to the adherend surface of the adherend. Therefore, the specific method for producing a laminate using the film-like coating agent can be the same as that for obtaining an adhesive using a film-like adhesive. When the softened film-like coating agent is pressed against the substrate surface to harden it, for example, the film-like coating agent may be pressed against the substrate surface using a pressing body similar to the pressing body used when pressing the solution-like coating agent coated on the substrate against the substrate surface. In addition, like the film-like adhesive, the film-like coating agent can be expected to have high adhesive strength to the substrate, and can also be peeled off after adhesion, and can be reused.

[0204] For the powdered adhesive or powdered coating agent containing a polymer compound, a powder composition containing a powder of the polymer compound obtained by the method of this embodiment is used. This powder composition may contain sub-components such as various residual additives as long as it contains a polymer compound as a main component. As described above, in the method for producing a polymer compound according to this embodiment, the target polymer compound is obtained in a state dissolved in dimethyl sulfoxide (as a dimethyl sulfoxide solution). Therefore, the method for producing a powdered adhesive or powdered coating agent according to this embodiment includes a step of powdering the polymer compound extracted from dimethyl sulfoxide by, for example, a known extraction method or separation method, by a known method.

[0205] The polymer compound contained in the powder adhesive or powder coating agent is not particularly limited as long as it is obtained by the manufacturing method according to this embodiment. For example, any of the polymer compounds contained in the solution adhesive or solution coating agent, the water-dispersible adhesive or water-dispersible coating agent, and the film adhesive or film coating agent may be used. As described above, the polymer compound according to this embodiment shows a higher affinity to an aqueous medium (aqueous liquid) containing water than proteins. Therefore, the powder adhesive or powder coating agent containing such a polymer compound can increase the protein content (dispersion amount) and can uniformly disperse the polymer compound in the aqueous medium compared to the water-dispersible adhesive or water-dispersible coating agent simply obtained by dispersing the protein in the aqueous medium. Therefore, by using the water-dispersible adhesive or water-dispersible coating agent in which the polymer compound is dispersed in the aqueous medium, the adhesive strength of the adhesive or coating agent to the adherend surface of the adherend or the laminate surface described later can be expected to be improved compared to the case of using the water-dispersible adhesive or water-dispersible coating agent in which the protein is dispersed in the aqueous medium.

[0206] A powdery adhesive containing a modified protein is also used to produce an adhesive in which a plurality of adherends are bonded to each other. For example, in a state where a powdery adhesive is interposed between a plurality of adherends to be bonded, the powdery adhesive is heated and pressurized through the adherends to solidify, thereby bonding the adherends together to obtain an adhesive. According to such a method, an adhesive is obtained using a biodegradable adhesive, and there is an advantage that an adhesive that can reduce the environmental load at the time of disposal can be easily produced. Note that the adherend may be any one that can be bonded with a powdery adhesive containing a polymer compound, and may be, for example, the same as an adherend that is bonded with a solution-based adhesive containing a polymer compound.

[0207] There is no particular limitation on the specific method for producing an adhesive using the powdery adhesive. For example, when heating and pressurizing the powdery adhesive interposed between a plurality of adherends to be bonded, the adherends are sandwiched between metal plates or the like from both sides opposite to the adherend surface side, and the metal plates are heated to heat the powdery adhesive interposed between the adherends together with the adherends. At the same time, the metal plate is pressed with a hand press or the like, and the powdery adhesive is pressed through the adherends for a predetermined time. This resinifies and solidifies the powdery adhesive to bond the adherends. The heating temperature, amount of pressure, and heating and pressing time for solidifying the powdery adhesive are appropriately selected from the ranges that can resinify the modified protein according to the type of modified protein contained in the powdery adhesive.

[0208] Powder coating agents containing polymer compounds are also used to manufacture laminates in which a coating layer is laminated on the entire or part of the surface of a substrate. For example, the powder coating agent is placed on at least a part of the surface of a substrate, and the powder coating agent is heated and pressed between a pressurizing body and the substrate to solidify, thereby laminating a coating layer on at least a part of the surface of the substrate.

[0209] There is no particular limitation on the specific method for producing a laminate using a powder coating agent. For example, a metal plate or the like is placed so as to cover the entire powder adhesive placed on the surface of the substrate, and the metal plate is heated to heat the powder adhesive. At the same time, the powder coating agent is resinified and solidified by pressing the powder adhesive between the metal plate and the substrate for a predetermined time using a hand press or the like. The heating temperature and amount of pressure, or the heating and pressing time, of the powder coating agent are the same as those used when obtaining an adhesive body using a powder adhesive.

[0210] Although several embodiments have been described above, the present disclosure is not limited to the above-described embodiments. In addition, the contents of the descriptions of the above-described embodiments can be mutually applied. EXAMPLES

[0211] The present disclosure will be described more specifically below based on examples, etc. However, the present disclosure is not limited to the following examples.

[0212] [Preparation of Polypeptides] (Preparation of expression vector) An artificial fibroin having the amino acid sequence shown in SEQ ID NO:9 (PRT2882) was designed.

[0213] The average hydropathic index value of the artificial fibroin having the amino acid sequence shown in SEQ ID NO:9 (PRT2882) is 0.47.

[0214] Nucleic acids encoding artificial fibroins having the amino acid sequence shown in SEQ ID NO:9 were synthesized. An NdeI site was added to the 5' end of the nucleic acid, and an EcoRI site was added downstream of the termination codon. The nucleic acids were cloned into a cloning vector (pUC118). The nucleic acids were then excised by restriction enzyme treatment with NdeI and EcoRI, and then recombined into a polypeptide expression vector pET-22b(+) to obtain an expression vector.

[0215] (Preparation of Polypeptides) The resulting expression vector was used to transform E. coli BLR(DE3). The transformed E. coli was cultured in 2 mL of LB medium containing ampicillin for 15 hours. The culture was transferred to 100 mL of seed culture medium (Table 2) containing ampicillin at OD 200. 600 The culture temperature was kept at 30°C, and the OD 600 The flask culture was continued until the number of cells reached 5 (about 15 hours), and a seed culture solution was obtained.

[0216] [Table 2]

[0217] The seed culture solution was transferred to a jar fermenter containing 500 mL of production medium (Table 3) and the OD 600 The culture temperature was kept at 37°C and the pH was controlled to be constant at 6.9. The dissolved oxygen concentration in the culture was maintained at 20% of the dissolved oxygen saturation concentration.

[0218] [Table 3]

[0219] Immediately after the glucose in the production medium was completely consumed, the feed solution (glucose 455g / 1L, yeast extract 120g / 1L) was added at a rate of 1mL / min. The culture temperature was kept at 37°C and the culture was controlled to a constant pH of 6.9. The dissolved oxygen concentration in the culture was maintained at 20% of the dissolved oxygen saturation concentration, and the culture was continued for 20 hours. Then, 1M isopropyl-β-thiogalactopyranoside (IPTG) was added to the culture solution to a final concentration of 1mM to induce the expression of artificial fibroin. After 20 hours had passed after the addition of IPTG, the culture solution was centrifuged and the cells were collected. SDS-PAGE was performed using the cells prepared from the culture solution before and after the addition of IPTG, and the expression of the desired artificial fibroin was confirmed by the appearance of a band of the desired artificial fibroin size depending on the addition of IPTG.

[0220] (Polypeptide purification) The cells harvested 20 hours after the addition of IPTG were washed with 20 mM Tris-HCl buffer (pH 7.4). The washed cells were suspended in 20 mM Tris-HCl buffer (pH 7.4) containing about 1 mM PMSF, and the cells were disrupted with a high-pressure homogenizer (GEA Niro Soavi). The disrupted cells were centrifuged to obtain a precipitate. The precipitate was washed with 20 mM Tris-HCl buffer (pH 7.4) until it reached a high purity. The washed precipitate was suspended in 8 M guanidine buffer (8 M guanidine hydrochloride, 10 mM sodium dihydrogen phosphate, 20 mM NaCl, 1 mM Tris-HCl, pH 7.0) to a concentration of 100 mg / mL, and stirred with a stirrer at 60°C for 30 minutes to dissolve. After dissolution, dialysis was performed against water using a dialysis tube (Cellulose tube 36 / 32, Sanko Junyaku Co., Ltd.).

[0221] The white aggregated polypeptide obtained after dialysis was collected by centrifugation, the water was removed using a freeze-dryer, and the freeze-dried powder was collected to obtain powdered artificial fibroin (PRT2882). It was confirmed that disulfide bonds were formed between molecules of the artificial fibroin into which cysteine ​​residues had been inserted. It was confirmed that the artificial fibroin contained 13.8 parts by mass of dimers, 2.1 parts by mass of trimers, and 0.8 parts by mass of tetramers per 100 parts by mass of monomers. The formation of disulfide bonds was measured by SDS-PAGE.

[0222] [Preparation of a compound having a structure represented by general formula (1)] (Synthesis of polyethylene glycol bismaleate) 500 g of polyethylene glycol (weight average molecular weight: 10000) and 50 g of maleic anhydride were placed in a container, and the container was evacuated and replaced with nitrogen. After the contents were melted, 7 mL of triethylamine was added, and the mixture was heated to 80° C. and stirred for 2.5 hours to react. A precipitate was generated and collected by a reprecipitation method in which the solution after the reaction was added to 500 mL of cyclopentyl methyl ether (CPME). 1500 mL of CPME was added again to the resulting precipitate, and the precipitate was washed by stirring. 2000 mL of tetrahydrofuran was added to the washed precipitate collected by filtration, and the precipitate was further washed by stirring. The precipitate was collected by filtration and dried in vacuum to obtain 450 g of polyethylene glycol bismaleate (polyethylene glycol having two maleic acid skeletons) (yield: 90% by mass).

[0223] [Preparation of a compound having a structure represented by formula (2)] (Synthesis of polyethylene glycol bisacrylate) 300 mL of a solution in which 60 g of polyethylene glycol (weight average molecular weight: 10,000) was dissolved in dichloromethane was prepared, and 60 g of anhydrous sodium sulfate was added and stirred for 30 minutes. The anhydrous sodium sulfate was collected by filtration, and the collected matter was washed with 100 mL of dichloromethane to remove solid matter, thereby obtaining a washing liquid. The obtained filtrate and washing liquid were placed in a 1 L four-neck flask, and the solvent was distilled off under reduced pressure to obtain a purified polyethylene glycol.

[0224] Next, under a nitrogen atmosphere, 300 mL of toluene that had been dehydrated in advance using molecular sieves 4A was added to the purified polyethylene glycol to prepare a suspension. The resulting suspension was heated to about 50°C under a nitrogen atmosphere to obtain a homogeneous solution. Toluene was distilled off from the resulting solution under reduced pressure. 300 mg of hydroquinone was added to the resulting residue as a polymerization inhibitor, and the mixture was dried under reduced pressure. Nitrogen gas was then filled into the four-neck flask.

[0225] Next, 500 mL of dichloromethane previously dehydrated with molecular sieves 4A was added to the four-neck flask using a cannula at room temperature under nitrogen flow. Furthermore, 8.4 mmL (60 mmol) of triethylamine previously dehydrated with molecular sieves 4A was added using a syringe at room temperature under nitrogen atmosphere to prepare a mixed solution. The mixed solution was maintained under a nitrogen atmosphere and cooled using an ice bath adjusted to -15°C. While adjusting the reaction temperature to -5°C to -3°C, 5 mL (60 mmol) of acrylic acid chloride was added dropwise under a nitrogen atmosphere over about 5 minutes. After the dropwise addition of acrylic acid chloride was completed, the temperature was naturally raised without removing the ice bath, and the mixture was stirred and reacted overnight under a nitrogen atmosphere.

[0226] The solvent was distilled off from the solution after the reaction under reduced pressure, and the resulting residue was washed with n-hexane. 600 mg of hydroquinone was added to the washed residue, and hot-extracted with 700 mL of tetrahydrofuran to obtain an extract. The extract was filtered using Celite 503, and 700 mL of n-hexane was added to the collected material to cause reprecipitation. The resulting precipitate was collected by filtration and dissolved in 500 mL of dichloromethane, to which 300 mg of hydroquinone was added, and the solvent was distilled off under reduced pressure. The resulting residue was dried under reduced pressure to obtain 58.8 g (yield: 99% by mass) of polyethylene glycol bisacrylate (polyether having two acrylate skeletons) as a pale yellow powder.

[0227] [Preparation of other compounds] (Synthesis of polyethylene glycol bismaleimide) Polyethylene glycol bismaleimide (Mal-PEG-Mal (Maleimide-PEG-Maleimide), molecular weight: 10,000) manufactured by Funakoshi Corporation was used.

[0228] (Example I-1) In a two-necked flask, 1 g (0.1 mmol) of artificial fibroin (polypeptide, 10 kDa) having the amino acid sequence (PRT2882) shown in SEQ ID NO: 9 prepared as described above and 12.5 mg (0.08 mmol) of dithiothreitol (DTT) as a reducing agent were added, and 12.5 g of dimethyl sulfoxide (DMSO) was added, and the mixture was heated and stirred at 70°C for 30 minutes to prepare solution A containing the polypeptide. The amount of the reducing agent was adjusted to 0.8 equivalents relative to the mercapto group of the polypeptide.

[0229] 1 g (0.1 mmol) of the polyethylene glycol bismaleate prepared as described above was weighed out and placed in a flask, and 7.5 g of dimethyl sulfoxide (DMSO, Fujifilm Wako Pure Chemical Industries, Ltd.) was added and dissolved by heating and stirring at 70° C. for 10 minutes. In this way, solution B containing polyethylene glycol bismaleate was prepared.

[0230] Solution A and solution B were mixed, and 110 μL (0.08 mmol) of triethylamine (TEA, Fujifilm Wako Pure Chemical Industries, Ltd.) was further added as a base to prepare a reaction solution. The amount of the polypeptide in the mixed solution was adjusted to 7 parts by mass relative to 100 parts by mass of dimethyl sulfoxide. The amount of the base was adjusted to 8 parts by mass based on the total mass of the polypeptide. Next, the reaction solution was heated and stirred at 70° C. for 90 minutes to allow the reaction to proceed. The solution after the reaction was centrifuged to remove the precipitate, and a solution containing a polymer compound in which a polypeptide portion and a polyethylene glycol portion were bound was obtained. The centrifugation was performed using a micro high-speed refrigerated centrifuge (Tomy Industrial Co., Ltd., product name: MX-307) at 15,000 rpm for 10 minutes.

[0231] <Evaluation of polymer compound manufacturing methods: Precipitates in the solution after reaction> The post-reaction solution before centrifugation was visually observed to confirm the presence or absence of precipitate. In addition, it was confirmed whether or not a gel-like substance was formed when the supernatant after removing the precipitate was centrifuged under the treatment conditions of 15,000 rpm for 10 minutes. The obtained results were evaluated according to the following criteria. The results are shown in Table 4. It was confirmed that the precipitate was mainly yellowish white and contained by-products generated by the formation of disulfide bonds between peptides. The gel-like substance was a brown, transparent solid. A: No precipitate was observed, and no gel-like matter was formed. B: Precipitation is observed, but no gel-like material is formed. C: No precipitate is observed, but a gel-like substance is formed. D: Precipitation is observed and a gel-like substance is also formed.

[0232] <Evaluation of polymer compound manufacturing methods: degree of conversion from raw materials to the desired product> The reaction solution prepared in Example I-1 was degassed by centrifugation, and 3 g of the degassed reaction solution was dropped and spread on a disposable tray (manufactured by AS ONE Corporation, product name: Balance Dish SCC, external dimensions 80 mm x 80 mm x 24 mm). Thereafter, the solution was pre-dried at 60°C for 8 hours, and further dried under reduced pressure at 80°C for 3 hours to evaporate the solvent, thereby producing a film with a thickness of 0.09 mm. 2 mg of the obtained film and 3.1 mg of tris(2-carboxyethyl)phosphine (TCEP) were dissolved in a TFANa-HFIP solution (1.3 mL) in which 85.3 mg of sodium trifluoroacetate (TFANa, manufactured by Fujifilm Wako Pure Chemical Industries, Ltd.) was dissolved in 1000 g of HFIP, to prepare a measurement sample. The measurement sample thus obtained was subjected to gel permeation chromatography measurement under the conditions described below. In the obtained chromatogram, the peak intensity (peak height) of the raw material polypeptide was set as P2, and the peak intensity (peak height) of the product polymer compound was set as P1, and the ratio (P1 / P2) was calculated. The higher this value, the more the reaction proceeded. The results are shown in Table 4.

[0233] The measurement conditions for the GPC measurement were as follows: Equipment: Agilent 1260 Infinity II liquid chromatography system (Agilent Technologies, Inc.) Detector: Agilent 1260 Infinity II refractive index detector (RID) (Agilent Technologies, Inc.) Column: 0.5 μm guard column filter (Shodex GPC HK-G, Showa Denko K.K.), styrene-divinylbenzene copolymer column (Showdex GPC HK404L x 2, inner diameter: 4.6 mm x length: 150 mm, Showa Denko K.K.) Eluent: TFANa-HFIP solution (85.3 mg of sodium trifluoroacetate (TFANa) dissolved in 1000 g of hexafluoroisopropanol (HFIP, Central Glass Co., Ltd.)) ·Flow rate: 0.15mL / min ·Measurement temperature: 40℃

[0234] (Example I-2) A polymer compound in which a polypeptide moiety and a polyethylene glycol moiety were bound was obtained in the same manner as in Example I-1, except that the components, their amounts, and the reaction temperature were adjusted as shown in Table 4.

[0235] (Example I-3) In a two-necked flask, 1 g (0.1 mmol) of artificial fibroin (polypeptide, 10 kDa) having the amino acid sequence (PRT2882) shown in SEQ ID NO: 9 prepared as described above and 7.2 mg (0.08 mmol) of butanediol as a reducing agent were added, and 9.0 g of dimethyl sulfoxide (DMSO) was added, and the mixture was heated and stirred at 70°C for 30 minutes to prepare solution C containing the polypeptide. The amount of the reducing agent was adjusted to 0.8 equivalents relative to the mercapto group of the polypeptide.

[0236] 1 g (0.1 mmol) of the polyethylene glycol bismaleate prepared as described above was weighed out and placed in a flask, and 3.0 g of dimethyl sulfoxide (DMSO, Fujifilm Wako Pure Chemical Industries, Ltd.) was added and dissolved by heating and stirring at 70° C. for 10 minutes. In this way, solution D containing polyethylene glycol bismaleate was prepared.

[0237] Solution C and solution D were mixed, and 110 μL (0.08 mmol) of triethylamine (TEA, Fujifilm Wako Pure Chemical Industries, Ltd.) was further added as a base to prepare a reaction solution. In the mixed solution, the amount of the polypeptide was adjusted to 7 parts by mass relative to 100 parts by mass of dimethyl sulfoxide. The amount of the base was adjusted to 8 parts by mass based on the total mass of the polypeptide. Next, the reaction solution was heated and stirred at 70° C. for 90 minutes to allow the reaction to proceed. The solution after the reaction was centrifuged to remove the precipitate, and a solution containing a polymer compound in which a polypeptide portion and a polyethylene glycol portion were bound was obtained. The centrifugation was performed using a micro high-speed refrigerated centrifuge (Tomy Industrial Co., Ltd., product name: MX-307) under the treatment conditions of 15,000 rpm for 10 minutes.

[0238] (Example I-4) A solution containing a polymer compound in which a polypeptide portion and a polyethylene glycol portion are bonded was obtained in the same manner as in Example I-2, except that the components, amounts thereof, and reaction temperature were adjusted as shown in Table 4.

[0239] (Example I-5) 1 g (0.1 mmol) of the polyethylene glycol bisacrylate prepared as described above and 25 mg (0.23 mmol) of hydroquinone (manufactured by FUJIFILM Wako Pure Chemical Industries, Ltd.) as a polymerization inhibitor were weighed into a flask, and 9.0 g of dimethyl sulfoxide (DMSO, manufactured by FUJIFILM Wako Pure Chemical Industries, Ltd.) was added and dissolved by heating and stirring at 70° C. for 10 minutes. In this way, a solution E containing polyethylene glycol bisacrylate was prepared.

[0240] Solution A prepared in the same manner as in Example I-1 was mixed with the above solution E, and 110 μL (0.08 mmol) of triethylamine (TEA, manufactured by Fujifilm Wako Pure Chemical Industries, Ltd.) was further added as a base to prepare a reaction solution. In the mixed solution, the polypeptide was adjusted to 7 parts by mass relative to 100 parts by mass of dimethyl sulfoxide. Next, the reaction solution was heated and stirred at 70° C. for 90 minutes to allow the reaction to proceed. The solution after the reaction was centrifuged to remove the precipitate, and a solution containing a polymer compound in which a polypeptide portion and a polyethylene glycol portion were bound was obtained. The centrifugation was performed using a micro high-speed refrigerated centrifuge (manufactured by Tomy Kogyo Co., Ltd., product name: MX-307) under the treatment conditions of 15,000 rpm for 10 minutes.

[0241] (Examples I-6 to I-7) A solution containing a polymer compound in which a polypeptide portion and a polyethylene glycol portion are bonded was obtained in the same manner as in Example I-5, except that the components, amounts thereof, and reaction temperature were adjusted as shown in Table 4.

[0242] In Examples I-2 to I-7, the solutions obtained after the reaction were evaluated in the same manner as in Example I-1. The results are shown in Table 4. [Table 4]

[0243] In Table 4, the amount of dimethyl sulfoxide means the total amount in the solution after mixing (for example, the sum of the amount of DMSO in solution A and the amount of DMSO in solution B).

[0244] (Comparative example I-1) 1 g (0.1 mmol) of polyethylene glycol bismaleimide (Mal-PEG-Mal (Maleimide-PEG-Maleimide), molecular weight 10000, manufactured by Funakoshi Co., Ltd.) was weighed out into a flask, and 12.5 g of dimethyl sulfoxide (DMSO, manufactured by Fujifilm Wako Pure Chemical Industries, Ltd.) was added and dissolved by heating and stirring at 70° C. for 10 minutes. In this way, solution F containing polyethylene glycol bismaleimide was prepared.

[0245] Solution A prepared in the same manner as in Example I-1 was mixed with the above solution F, and 110 μL (0.08 mmol) of triethylamine (TEA, manufactured by Fujifilm Wako Pure Chemical Industries, Ltd.) was further added as a base to prepare a reaction solution. In the mixed solution, the amount of the polypeptide was adjusted to 7 parts by mass relative to 100 parts by mass of dimethyl sulfoxide. Next, the reaction solution was heated and stirred at 70° C. for 90 minutes to allow the reaction to proceed. The solution after the reaction was centrifuged to obtain a polymer compound in which a polypeptide portion and a polyethylene glycol portion were bound as a precipitate. The centrifugation was performed using a micro high-speed refrigerated centrifuge (manufactured by Tomy Kogyo Co., Ltd., product name: MX-307) at 15,000 rpm for 10 minutes.

[0246] (Comparative example I-2) In a two-necked flask, 1 g (0.1 mmol) of artificial fibroin (polypeptide, 10 kDa) having the amino acid sequence (PRT2882) shown in SEQ ID NO: 9 prepared as described above and 12.5 mg (0.08 mmol) of dithiothreitol (DTT) as a reducing agent were added, and 12.5 g of dimethyl sulfoxide (DMSO) was added, and the mixture was heated and stirred at 70°C for 30 minutes to prepare solution G containing the polypeptide. The amount of the reducing agent was adjusted to 0.8 equivalents relative to the mercapto group of the polypeptide.

[0247] 1 g (0.1 mmol) of polyethylene glycol bismaleimide (Mal-PEG-Mal (Maleimide-PEG-Maleimide), molecular weight 10,000, manufactured by Funakoshi Co., Ltd.) was weighed out into a flask, and 25.0 g of dimethyl sulfoxide (DMSO, manufactured by Fujifilm Wako Pure Chemical Industries, Ltd.) was added and dissolved by heating and stirring at 70° C. for 10 minutes. In this way, solution H containing polyethylene glycol bismaleimide was prepared.

[0248] The above solution G and the above solution H were mixed, and 110 μL (0.08 mmol) of triethylamine (TEA, manufactured by Fujifilm Wako Pure Chemical Industries, Ltd.) was further added as a base to prepare a reaction solution. In the mixed solution, the amount of the polypeptide was adjusted to 7 parts by mass relative to 100 parts by mass of dimethyl sulfoxide. Next, the reaction solution was heated and stirred at 70° C. for 90 minutes to allow the reaction to proceed. The solution after the reaction was centrifuged to obtain a polymer compound in which a polypeptide portion and a polyethylene glycol portion were bound as a precipitate. The centrifugation was performed using a micro high-speed refrigerated centrifuge (manufactured by Tomy Industrial Co., Ltd., product name: MX-307) at 15,000 rpm for 10 minutes.

[0249] In Comparative Examples I-1 and I-2, the resulting post-reaction solutions were evaluated in the same manner as in Example I-1. The results are shown in Table 5. In Table 5, the peak intensity ratio in GPC for Comparative Example 1 is marked "-", which indicates that the measurement itself was not possible due to precipitation and gel-like matter in the reaction system.

[0250] [Table 5]

[0251] (Example I-8) The reaction solution prepared in Example I-3 was degassed by centrifugation, and 3 g of the degassed reaction solution was dropped and spread on a disposable tray (As One Corporation, product name: Balance Dish SCC, external dimensions 80 mm x 80 mm x 24 mm). Thereafter, the solution was dried at 60°C for 8 hours, and further dried under reduced pressure at 80°C for 3 hours to evaporate the solvent, thereby producing a film with a thickness of 0.09 mm.

[0252] <Film evaluation> The film prepared as described above was measured for breaking elongation, elastic modulus, and toughness using a tensile tester (manufactured by Shimadzu Corporation, product name: AG-X plus50kN). The film was stored for 24 hours or more under an environment of 20°C and relative humidity: 65% before use. After storage, a dumbbell-shaped test specimen was prepared by punching out the film at the measurement site with a film distance of 25 mm and a width of 10 mm using a punching machine (manufactured by Tester Sangyo Co., Ltd., product name: SA-1008). The test specimen was set in the tensile tester, and a tensile test was performed under the conditions of a load cell of 50 kN, a gripper distance of 25 mm, and a tensile speed of 10 mm / min. The results are shown in Table 6.

[0253] (Example I-9) A film was prepared in the same manner as in Example I-8, except that the polymer material was changed to that prepared in Example I-7. The obtained film was evaluated in the same manner as in Example I-8. The results are shown in Table 6.

[0254] (Comparative example I-3) A film was prepared in the same manner as in Example I-8, except that the polymer material was changed to that prepared in Comparative Example I-2. The obtained film was evaluated in the same manner as in Example I-8. The results are shown in Table 6.

[0255] [Table 6]

[0256] As shown in Table 6 above, since side reactions such as the formation of disulfide bonds between polypeptides were suppressed, no precipitate was formed, and even though the solutions containing the polymers prepared in Examples I-8 and I-9 were used to form films without purification, it was confirmed that films exhibiting good physical properties could be obtained.

[0257] (Example I-10) 100 g of the solution prepared in the same manner as in Example I-1 was gradually dropped into 100 g of ethyl acetate prepared in a 300 mL beaker while stirring. The precipitate formed in the beaker was finely pulverized using a homogenizer (manufactured by IKA, product name: T-18 Digital). The precipitate was then collected by suction filtration. The collected precipitate was put into 100 g of ethyl acetate, stirred and washed. After repeating washing with ethyl acetate twice, the precipitate was collected by filtration and vacuum dried at 40° C. for 2 hours to obtain a powder of a purified polymer compound.

[0258] 6 g of the obtained powder was added to a 20 mL glass vial (As One, product name: screw vial) containing a stirrer, and then 11.46 mL of formic acid (Niwagosei Co., Ltd., concentration: 98% by mass) was added. The mixture was then stirred at 45°C for 2 hours to prepare a formic acid solution (dope solution) with a concentration of 30% by mass. The viscosity of the obtained dope solution at 45°C was 17,300 mPa·s. The viscosity of the dope solution was measured by filling the dope solution into a glass tube containing a spherical probe (φ4.7 mm) using an EMS viscometer (Kyoto Electronics Manufacturing Co., Ltd., product name: EMS-1000S) at a measurement temperature of 45°C.

[0259] 10.0 mL of the above dope solution was filled into a 50 mL disposable syringe (Musashi Engineering, product name: PSY-50E), and then a plunger (Musashi Engineering, product name: FLP-50E) was inserted. A plastic needle (Musashi Engineering, product name: 22, inner diameter: 0.47 mm) was attached, and an adapter tube (Musashi Engineering, product name: AT-50E-H-1.0) was attached to the upper part of the syringe. A syringe heater was also attached, and the dope solution was kept at 45°C.

[0260] Then, the dope solution was discharged at a linear velocity of 1.00 m / min by operating a syringe pump kept at 45° C. while flowing in 0.2 MPa of nitrogen. The discharged dope solution was passed through a drying line at 200° C., and then passed through a take-up roller adjusted to a rotation speed of 10 m / min.

[0261] Then, the fiber was passed through a drying line at 275°C via two rollers and wound up by a winder with a rotation speed of 15 m / min to obtain a fiber (filament). The mass per meter of the wound fiber was measured, and the mass per 10,000 m (fineness, unit: dtex) was calculated. A sample was prepared by bundling five of the filaments, and a cross-sectional analysis was performed using a polarizing microscope (manufactured by Nikon Corporation, product name: ECLIPSE LV100ND) to determine the circle-equivalent diameter of the five filaments, and the arithmetic average was adopted as the circle-equivalent diameter of the filament.

[0262] <Fiber evaluation: Measurement of breaking elongation and breaking strength> The breaking elongation and breaking strength of the sample obtained by bundling five of the filaments were evaluated using a tensile tester (AG-X plus50kN, manufactured by Shimadzu Corporation). The sample was stored for 24 hours or more in an environment of 20°C and 65% humidity before use. The sample was set in the tester and stretched until it broke under the following conditions: load cell: 50kN, gripper distance: 50mm, and tensile speed: 50cm / min. The stress and length at the time of break were measured three times, and the arithmetic average value was calculated. The results are shown in Table 7.

[0263] <Fiber evaluation: Stress relaxation measurement> The stress relaxation of the sample obtained by bundling the five filaments was evaluated using a tensile tester (manufactured by Shimadzu Corporation, product name: AG-X plus50kN). The sample was stored for 24 hours or more in an environment of 20°C and early-stage humidity: 65% before use. The sample was set in the tester, and a tensile test was performed under the conditions of a load cell of 50kN, a distance between grippers of 50mm, and a tensile speed of 50cm / min. At this time, the stress X was measured when the 50mm sample was stretched 300%. After that, the sample was held in that state for 30 seconds, and the stress Y immediately after the holding was measured. This measurement was performed three times, and the stress relaxation was calculated using the formula (XY) / X×100, and the arithmetic average value was taken as the stress relaxation. The results are shown in Table 7.

[0264] (Reference example) A commercially available urethane fiber (manufactured by Toray Opelontex Co., Ltd., product name: Lycra T-127C) was evaluated in the same manner as in Example I-10. The results are shown in Table 7.

[0265] [Table 7]

[0266] (Example II-1) To 3.13 g of a solution containing a polymer compound in which a polypeptide portion and a polyethylene glycol portion are bonded, prepared in the same manner as in Example I-1, 50 mg of a water-soluble thermoplastic resin (manufactured by Meisei Chemical Industry Co., Ltd., product name: Alkox E-75) was added, and the mixture was stirred at 70°C for 1 hour to prepare a DMSO solution (dope solution). The viscosity of the obtained dope solution at a temperature of 25°C was 4130 mPa·s. The viscosity of the dope solution here was a value measured at a measurement temperature of 25°C using a digital viscometer (manufactured by Ametech Co., Ltd., product name: Brookfield DV2TLVTJ0).

[0267] The dope was transferred to a 100 mL glass beaker. The dope was slowly stirred with a stainless steel spatula to avoid air bubbles, and the dope was slowly lifted up to stretch it into a fiber shape by utilizing its viscosity. The stretched fiber-like dope was placed on a release film and vacuum-dried for 15 hours in a vacuum oven at 80°C to obtain a filament (a polymer compound formed into a fiber shape).

[0268] A sample was prepared by bundling five of the above filaments, and cross-sectional analysis was performed using a polarizing microscope (Nikon Corporation, product name: ECLIPSE LV100ND) to determine the circular equivalent diameters of the five filaments, and the arithmetic mean was adopted as the circular equivalent diameter of the filament.

[0269] The five resulting filaments were bundled into a sample, and the fiber was evaluated in the same manner as in Example I-10. The results are shown in Table 8. In Table 8, "-" indicates that no measurement was performed.

[0270] [Table 8]

[0271] (Comparative example I-4) An attempt was made to prepare fibers by changing the polymer material to that prepared in Comparative Example I-2, but the concentration of the solution was insufficient and fibers could not be produced.

[0272] (Examples III-1 to III-4) A solution containing a polymer compound in which a polypeptide portion and a polyethylene glycol portion are bonded was obtained in the same manner as in Example I-1, except that the components, amounts thereof, and reaction temperature were adjusted as shown in Table 9.

[0273] In Examples III-1 to III-4, the solutions obtained after the reaction were evaluated in the same manner as in Example I-1. The results are shown in Table 9.

[0274] [Table 9]

[0275] (Examples III-5 to III-6) A solution containing a polymer compound in which a polypeptide portion and a polyethylene glycol portion are bonded was obtained in the same manner as in Example I-1, except that the components, amounts thereof, and reaction temperature were adjusted as shown in Table 10.

[0276] In Examples III-5 and III-6, the solutions obtained after the reaction were evaluated in the same manner as in Example I-1. The results are shown in Table 10.

[0277] (Comparative Example III-1, Examples III-7 to III-8) A solution containing a polymer compound in which a polypeptide portion and a polyethylene glycol portion are bonded was obtained in the same manner as in Example I-1, except that the components, amounts thereof, and reaction temperature were adjusted as shown in Table 10.

[0278] In Comparative Example III-1 and Examples III-7 to III-8, the solutions obtained after the reaction were evaluated in the same manner as in Example I-1. The results are shown in Table 10.

[0279] [Table 10]

[0280] (Examples III-9 to III-11) A solution containing a polymer compound in which a polypeptide portion and a polyethylene glycol portion are bonded was obtained in the same manner as in Example I-1, except that the components, amounts thereof, and reaction temperature were adjusted as shown in Table 11.

[0281] In Examples III-9 to III-11, the solutions obtained after the reaction were evaluated in the same manner as in Example I-1. The results are shown in Table 11. For reference, Table 11 also shows the results of Examples I-1 and III-2.

[0282] [Table 11]

[0283] (Example I-11) <Production of film adhesive> In a two-necked flask, 2.5 g (0.25 mmol) of artificial fibroin (polypeptide, 10 kDa) having the amino acid sequence (PRT2882) shown in SEQ ID NO: 9 prepared as described above and 31.2 mg (0.2 mmol) of dithiothreitol (DTT) as a reducing agent were added, and 25 g of dimethyl sulfoxide (DMSO) was added, and the mixture was heated and stirred at 70°C for 30 minutes to prepare solution I containing the polypeptide. The amount of the reducing agent was adjusted to 0.8 equivalents relative to the mercapto group of the polypeptide.

[0284] 2.5 g (0.025 mmol) of the polyethylene glycol bismaleate prepared as described above was weighed out and placed in a flask, and 20 g of dimethyl sulfoxide (DMSO, Fujifilm Wako Pure Chemical Industries, Ltd.) was added, and the mixture was heated and stirred at 70 for 10 minutes to dissolve the polyethylene glycol bismaleate. In this way, a solution J containing polyethylene glycol bismaleate was prepared.

[0285] Solution I and solution J were mixed, and 275 μL (0.2 mmol) of triethylamine (TEA, Fujifilm Wako Pure Chemical Industries, Ltd.) was further added as a base to prepare a reaction solution. In the mixed solution, the amount of the polypeptide was adjusted to 6 parts by mass relative to 100 parts by mass of dimethyl sulfoxide. The amount of the base was adjusted to 8 parts by mass based on the total mass of the polypeptide. Next, the reaction solution was heated and stirred at 70° C. for 90 minutes to allow the reaction to proceed. The solution after the reaction was centrifuged to degas it, and a DMSO solution containing a polymer compound in which a polypeptide portion and a polyethylene glycol portion were bound was obtained. The centrifugation was performed using a micro high-speed refrigerated centrifuge (Tomy Industrial Co., Ltd., product name: MX-307) at 15,000 rpm for 10 minutes.

[0286] Next, the DMSO solution of the obtained polymer compound was dropped onto a stainless steel plate covered with a PET sheet (Teijin Film Solutions Co., Ltd., Purex, A54 type, thickness 38 μm) and spread using a frame applicator with an analog micrometer (Imoto Manufacturing Co., Ltd., 250 mm coating width, 1 mm gap) and a coating machine (Imoto Manufacturing Co., Ltd., IMC-7370, coating speed 10 mm / 1 sec). Next, it was dried at 60 ° C for 8 hours, and further dried under reduced pressure at 80 ° C for 3 hours to evaporate the DMSO, obtaining a film-like adhesive with a thickness of 0.08 mm.

[0287] <Production of Adhesive Body> The film-like adhesive obtained as described above was cut to 15 mm x 15 mm, and the cut film-like adhesive was immersed in water for 5 minutes to swell and soften. Next, the softened film-like adhesive was placed on one end of a wooden board (made of cypress, 1.5 mm x 40 mm x 2 mm) in an area of ​​15 mm x 15 mm, and another wooden board of the same size was placed on top of the two wooden boards so that the film-like adhesive was interposed between them. After that, the two wooden boards were fixed with clips so that the film-like adhesive was pressed against each wooden board, and placed in a constant temperature thermostat with a blower and dried at 60 ° C for 1 hour to harden the film-like adhesive. In this way, an adhesive body A in which two wooden boards were bonded with a film-like adhesive was obtained. For comparison, an adhesive body B in which two pieces of wood were bonded with a double-sided tape was obtained by carrying out the same operation as above, except that a commercially available double-sided tape (Nichiban, 15 mm wide, Nicetack NW-15) was used instead of the film-like adhesive.

[0288] <Tensile test of adhesive bond> The adhesive body A and the adhesive body B obtained as described above were each set in a tensile tester (AG-X plus50kN, manufactured by Shimadzu Corporation) at room temperature of 20°C and humidity of 65% and subjected to a tensile test. The tensile test was performed under the following conditions: load cell 50kN, gripping distance 57.5mm, and tensile speed 10mm / min. As a result, the tensile strength of the adhesive body A was more than 10 times greater than that of the adhesive body B. This demonstrated that the film-like adhesive has a sufficiently large adhesive force (adhesive strength) compared to commercially available double-sided tapes and the like.

[0289] <Peel test of adhesive> Another bonded body A was produced in the manner described above, and the film-like adhesive that had protruded from the gap in the bonded portion of this bonded body A was pulled out from the bonded portion. This enabled the two bonded wooden boards to be peeled off. Furthermore, when the surfaces of the bonded portions of the two wooden boards were visually observed, no change was observed between the surfaces of the wooden boards before and after bonding. Furthermore, after the pulled-out film-like adhesive was washed with water, the same operation as that for producing the bonded body A was carried out using this film-like adhesive and two other wooden boards. As a result, two other wooden boards could also be bonded. This shows that the film-like adhesive can be reused.

[0290] (Example I-12) <Production of powder adhesive> 319.3 g of dimethyl sulfoxide (DMSO) was measured into a three-necked separable flask, and 419 mg (2.72 mmol) of dithiothreitol (DTT) was added as a reducing agent and stirred to dissolve. Then, 27.5 g (2.75 mmol) of artificial fibroin (polypeptide, 10 kDa) having the amino acid sequence (PRT2882) shown in SEQ ID NO: 9 prepared as described above was added, and the mixture was heated and stirred at 70°C for 30 minutes to prepare solution K containing the polypeptide. The amount of the reducing agent was adjusted so that it was approximately 1 equivalent to the mercapto group of the polypeptide.

[0291] 27.5 g (2.75 mmol) of the polyethylene glycol bismaleate prepared as described above was weighed out and placed in a flask, and 125.5 g of dimethyl sulfoxide (DMSO, Fujifilm Wako Pure Chemical Industries, Ltd.) was added, and the mixture was heated and stirred at 70 for 10 minutes to dissolve the mixture. In this way, a solution L containing polyethylene glycol bismaleate was prepared.

[0292] Solution K and solution L were mixed, and 4400 mg (43.5 mmol) of triethylamine (TEA, Fujifilm Wako Pure Chemical Industries, Ltd.) was further added as a base to prepare a reaction solution. In the mixed solution, the amount of the polypeptide was adjusted to 5.5 parts by mass relative to 100 parts by mass of dimethyl sulfoxide. The amount of the base was adjusted to 16 parts by mass based on the total mass of the polypeptide. Next, the reaction solution was heated and stirred at 70°C for 90 minutes to allow the reaction to proceed. The solution after the reaction was centrifuged to remove the precipitate, and a DMSO solution containing a polymer compound in which a polypeptide portion and a polyethylene glycol portion were bound was obtained. The centrifugation was performed at 15,000 rpm for 10 minutes.

[0293] The obtained DMSO solution of the polymer compound was washed in acetone and centrifuged five times, after which the supernatant was removed by decantation to separate the precipitate. The obtained precipitate was spread on a tray and dried in a fume hood for several hours, and then dried overnight in a vacuum oven at 40°C to obtain a powder of the polymer compound. The residual DMSO in this powder was less than 3% by mass. The resulting polymer powder was used as a powder adhesive.

[0294] <Manufacture of Laminate> 0.5 g of the powdery adhesive was weighed out and placed on a portion of one side of a cotton woven fabric substrate. The substrate with the powdery adhesive placed thereon was then sandwiched between two stainless steel plates coated with a release agent, and then heated to 100°C and pressed with a hand press at 10 MPa for 3 minutes. As a result, the powdery adhesive was resinified and solidified. This resulted in a laminate in which a coating layer was formed on a portion of the substrate surface. [Industrial Applicability]

[0295] According to the present disclosure, in the production of the above-mentioned polymer compound, a production method capable of increasing the raw material conversion rate compared to conventional methods can be provided.

Claims

1. A polypeptide having at least one mercapto group; At least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, having two structures represented by the following general formula (1): 【Chemistry 1】 [In the general formula (1), M is H, Na, K, NHEt ３ , or NHEtiPr ２ Either in dimethyl sulfoxide in the presence of a base and a reducing agent,

2. A polypeptide having at least one mercapto group; At least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, which has two structures represented by the following general formula (2): 【Chemistry 2】 [In general formula (2), R １ represents a hydrogen atom or a methyl group. in dimethyl sulfoxide in the presence of a base, a reducing agent and a polymerization inhibitor,

3. A polypeptide having at least one mercapto group; At least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, having two structures represented by the following general formula (1): 【Chemistry 3】 [In the general formula (1), M is H, Na, K, NHEt ３ , or NHEtiPr ２ Either in dimethyl sulfoxide in the presence of a reducing agent, by heating and reacting the above.

4. A polypeptide having at least one mercapto group; At least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, which has two structures represented by the following general formula (2): 【Chemistry 4】 [In general formula (2), R １ represents a hydrogen atom or a methyl group. in dimethyl sulfoxide in the presence of a reducing agent and a polymerization inhibitor,

5. The method for producing a polymer compound according to any one of claims 1 to 4, wherein the reducing agent is at least one selected from the group consisting of dithiol, sodium sulfate, sodium sulfite, and sodium dithionite.

6. The method according to any one of claims 1 to 4, wherein the amount of the reducing agent is 0.5 equivalents or more relative to the mercapto group of the polypeptide.

7. The method according to any one of claims 1 to 4, wherein the reaction between the polypeptide and the compound is carried out at 50°C or higher.

8. A method for producing a polymer solution, comprising dissolving the polymer obtained by the method according to any one of claims 1 to 4 in a solvent.

9. A method for producing a film, comprising forming a film from the polymer solution obtained by the method according to claim 8 .

10. A method for producing fibers, comprising spinning a polymer solution obtained by the method according to claim 8.

11. A method for producing a polymer compound solution by reacting a polypeptide having at least one mercapto group with at least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, each of which has two structures represented by the following general formula (1), in dimethyl sulfoxide in the presence of a base and a reducing agent under heating; and forming a film from the polymer compound solution. 【Chemistry 5】 [In the general formula (1), M is H, Na, K, NHEt ３ , or NHEtiPr ２ Either of the following is indicated.]

12. A method for producing a polymer compound solution by reacting a polypeptide having at least one mercapto group with at least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, the compound having two structures represented by the following general formula (2), in dimethyl sulfoxide in the presence of a base, a reducing agent, and a polymerization inhibitor, under heating; and forming a film from the polymer compound solution. 【Chemistry 6】 [In general formula (2), R １ represents a hydrogen atom or a methyl group.

13. A method for producing a polymer compound solution by reacting a polypeptide having at least one mercapto group with at least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, each of which has two structures represented by the following general formula (1), in dimethyl sulfoxide in the presence of a base and a reducing agent under heating; and spinning the polymer solution. 【Chemistry 7】 [In the general formula (1), M is H, Na, K, NHEt ３ , or NHEtiPr ２ Either of the following is indicated.]

14. A method for producing a polymer compound solution by reacting a polypeptide having at least one mercapto group with at least one compound selected from the group consisting of polyethers, polyesters, and polycarbonates, the compound having two structures represented by the following general formula (2), in dimethyl sulfoxide in the presence of a base, a reducing agent, and a polymerization inhibitor, under heating; and spinning the polymer solution. 【Chemistry 8】 [In general formula (2), R １ represents a hydrogen atom or a methyl group.

15. A film comprising a polymer compound, The polymer compound is A polypeptide portion; and at least one structural unit selected from the group consisting of a polyether structure, a polyester structure, and a polycarbonate structure, wherein the polypeptide portion is directly bound to S of the following general formula (3), (4), or (5) via a structure represented by the following general formula (3), (4), or (5): 【Chemistry 9】 【Chemistry 10】 【Chemistry 11】 [In the general formula (3) and the general formula (4), M is H, Na, K, NHEt ３ , or NHEtiPr ２ In the general formula (5), R １ represents H or Me. The elongation is 400% or more, A film having a ratio of break strength to yield strength greater than 1.

16. A fiber comprising a polymer compound, The polymer compound is A polypeptide portion; and at least one structural unit selected from the group consisting of a polyether structure, a polyester structure, and a polycarbonate structure, wherein the polypeptide portion is directly bound to S of the following general formula (3), (4), or (5) via a structure represented by the following general formula (3), (4), or (5): 【Chemistry 12】 【Chemistry 13】 【Chemistry 14】 [In the general formula (3) and the general formula (4), M is H, Na, K, NHEt ３ , or NHEtiPr ２ In the general formula (5), R １ represents H or Me. The elongation is 400% or more, A fiber having a ratio of breaking strength to yield strength greater than 1.

17. A method for producing a solution-type adhesive, comprising a step of dissolving the polymer compound obtained by the method according to any one of claims 1 to 4 in a solvent.

18. A method for producing a water-dispersible adhesive, comprising a step of dispersing the polymer compound obtained by the method according to any one of claims 1 to 4 in an aqueous medium.

19. A method for producing a film-like adhesive, comprising a step of forming a film from a solution in which a polymer compound obtained by the method according to any one of claims 1 to 4 is dissolved.

20. A method for producing a powdery adhesive, comprising the step of obtaining a powder composition containing the polymer compound obtained by the method according to any one of claims 1 to 4.

21. A method for producing an adherend, comprising: placing a solution obtained by dissolving a polymer compound obtained by the method according to any one of claims 1 to 4 in a solvent between a plurality of adherends; removing the solvent from the solution; and solidifying the polymer compound, thereby adhering the adherends to each other.

22. A method for producing an adherend, comprising: interposing an aqueous dispersion in which a polymer compound obtained by the method according to any one of claims 1 to 4 is dispersed in an aqueous medium between a plurality of adherends; removing the aqueous medium from the aqueous dispersion; and solidifying the polymer compound, thereby adhering the adherends to each other.

23. A method for producing an adhesive, comprising the steps of: softening a film containing a polymer compound obtained by the method according to any one of claims 1 to 4 by swelling or heating; interposing the film between a plurality of adherends; and curing the film while the film is pressed against the adherends, thereby bonding the adherends together.

24. 5. A method for producing an adherend, comprising: heating a powder composition containing a polymer compound obtained by the method according to any one of claims 1 to 4 in a state in which the powder composition is interposed between a plurality of adherends; and applying pressure to the powder composition via the adherends to solidify the powder composition, thereby adhering the adherends to each other.

25. A method for producing a solution-type coating agent, comprising a step of dissolving the polymer compound obtained by the method according to any one of claims 1 to 4 in a solvent.

26. A method for producing a water-dispersible coating agent, comprising a step of dispersing the polymer compound obtained by the method according to any one of claims 1 to 4 in an aqueous medium.

27. A method for producing a film-like coating agent, comprising a step of forming a film from a solution in which a polymer compound obtained by the method according to any one of claims 1 to 4 is dissolved.

28. A method for producing a powder coating agent, comprising the step of obtaining a powder composition containing the polymer compound obtained by the method according to any one of claims 1 to 4.

29. A method for producing a laminate having a substrate and a coating layer formed on at least a portion of a surface of the substrate, the method comprising the steps of: A method for producing a laminate, comprising: supplying a solution in which a polymer compound obtained by the method according to any one of claims 1 to 4 is dissolved in a solvent to at least a part of a surface of the substrate; coating at least a part of the surface of the substrate with the solution; and removing the solvent from the solution to solidify the polymer compound, thereby forming a coating layer on at least a part of the surface of the substrate.

30. A method for producing a laminate having a substrate and a coating layer formed on at least a portion of a surface of the substrate, the method comprising the steps of: A method for producing a laminate, comprising: supplying an aqueous dispersion in which a polymer compound obtained by the method according to any one of claims 1 to 4 is dispersed in an aqueous medium to at least a part of a surface of the substrate; coating at least a part of the surface of the substrate with the aqueous dispersion; and removing the aqueous medium from the aqueous dispersion to solidify the polymer compound, thereby forming a coating layer on at least a part of the surface of the substrate.

31. A method for producing a laminate having a substrate and a coating layer formed on at least a portion of a surface of the substrate, the method comprising the steps of: A method for producing a laminate, comprising the steps of: softening a film containing a polymer compound obtained by the method according to any one of claims 1 to 4 by swelling or heating; placing the film on at least a part of a surface of the substrate; and curing the film in a state of being pressed against the substrate, thereby laminating and forming the coating layer on at least a part of the surface of the substrate.

32. A method for producing a laminate having a substrate and a coating layer formed on at least a portion of a surface of the substrate, the method comprising the steps of:

5. A method for producing a laminate, comprising: placing a powder composition containing a polymer compound obtained by the method according to any one of claims 1 to 4 on at least a part of a surface of the substrate; heating the powder composition while pressing the powder composition between a pressurizing body and the substrate to solidify the powder composition, thereby forming the coating layer on at least a part of the surface of the substrate.

Images