109 results about "Single mutation" patented technology

The invention discloses a mutant protein screening method and device and related equipment thereof, and relates to the technical field of material determination and analysis. The method comprises the following steps: respectively carrying out simulated single mutation on a plurality of amino acid sites in a protein sequence of a target protein to obtain a plurality of simulated mutant proteins, and determining adaptability evaluation values F of the plurality of simulated mutant proteins based on a target protein language model; determining structural stability values S of the plurality of simulated mutant proteins based on a protein folding energy calculation tool; performing normalization processing on the adaptability evaluation value F and the structural stability value S, and performing single-index sorting and comprehensive-index sorting on normalization processing results; and screening the plurality of simulated mutant proteins according to the target screening number, the single index sorting result and the comprehensive index sorting result to obtain a target mutant protein. According to the method, the protein design transformation difficulty can be reduced, the mutant protein screening effect is improved, the calculation amount required by protein transformation and screening is reduced, and the consumption cost is reduced.

The invention belongs to the field of biological pharmacy and biotransformation, and particularly relates to a tagatose-4-epimerase mutant and application thereof. The tagatose-4-epimerase mutant disclosed by the invention is obtained by carrying out single mutation or combined mutation on an amino acid residue at a specific position of an amino acid sequence as shown in SEQ ID NO.1. Compared with wild type tagatose-4-epimerase, the mutant has the advantage that the catalytic activity is improved when the D-tagatose is prepared through conversion. The obtained tagatose-4-epimerase mutant has the advantages of being mild in reaction condition, green, environmentally friendly, single in product and easy to separate and purify, and the production cost is greatly reduced.

The invention discloses a streptomyces-derived cytochrome P450 enzyme mutant and an application of the streptomyces-derived cytochrome P450 enzyme mutant in synthesis of ursodesoxycholic acid. The cytochrome P450 enzyme mutant is obtained by carrying out single mutation or multiple mutation on the 181st site, the 192nd site, the 249th site or the 288th site of an amino acid sequence shown in SEQ ID NO.2. The cytochrome P450 enzyme mutant has high activity and regioselectivity, low-cost lithocholic acid is used as a substrate, efficient cytochrome P450 enzyme is used for hydroxylating the 7beta site of the lithocholic acid, ursodesoxycholic acid is synthesized in one step, the industrial production cost is greatly reduced, and the cytochrome P450 enzyme mutant has good application prospects in industrial production of ursodesoxycholic acid.

The invention discloses a 3-sterone-delta1-dehydrogenase mutant and application of the 3-sterone-delta1-dehydrogenase mutant in preparation of a steroid drug intermediate. The mutant is obtained by performing single mutation or multiple mutation on 551, 468, 48 and 157 sites of an amino acid sequence of 3-sterone-delta1-dehydrogenase as shown in SEQ ID NO: 1. The 3-ketosterone-delta1-dehydrogenase is subjected to molecular modification through directed evolution and semi-rational design, the mutant with improved enzyme activity is screened in a high-throughput mode, the mutant can effectively improve the conversion rate of a substrate 11alpha, 17alpha-dihydroxyprogesterone, and the substrate conversion rate of the mutant ISM-2 is improved to 92.4% compared with 36.7% before mutation.

The invention discloses an L-aspartic acid transaminase mutant and application thereof. The L-aspartic acid transaminase mutant is obtained by performing single mutation on the 104th amino acid of the amino acid sequence of L-aspartic acid transaminase. The L-aspartic acid transaminase mutant is used for synthesizing an L-phenylglycine derivative. The L-aspartic acid transaminase mutant is from escherichia coli, homologous sequence analysis and molecular docking simulation are carried out, site-specific mutagenesis verification is combined, and single-point mutation is carried out on the 104 amino acid to obtain the T104S mutant. The L-aspartic acid transaminase mutant disclosed by the invention can be used for efficiently catalyzing to generate an L-phenylglycine derivative, and compared with a wild type, the specific activity of the T104S mutant to o-chlorophenoxyacetic acid is 3.2 times that of the wild type.

The invention belongs to the field of biological pharmacy and biotransformation, and particularly relates to a tagatose-4-epimerase mutant, a coding gene and application. The tagatose-4-epimerase mutant disclosed by the invention is obtained by carrying out single mutation or combined mutation on an amino acid residue at a specific position of an amino acid sequence as shown in SEQ ID NO.1. Compared with wild type tagatose-4-epimerase, the mutant has the advantage that the catalytic activity is improved when the D-tagatose is prepared through conversion. The obtained tagatose-4-epimerase mutant has the advantages of being mild in reaction condition, green, environmentally friendly, single in product and easy to separate and purify, and the production cost is greatly reduced.

The invention belongs to the field of biological pharmacy and biotransformation, and particularly relates to a tagatose-4-epimerase mutant as well as a coding gene and application thereof. The tagatose-4-epimerase mutant disclosed by the invention is obtained by carrying out single mutation or combined mutation on an amino acid residue at a specific position of an amino acid sequence as shown in SEQ ID NO.1. Compared with wild type tagatose-4-epimerase, the mutant has the advantage that the catalytic activity is improved when the D-tagatose is prepared through conversion. The obtained tagatose-4-epimerase mutant has the advantages of being mild in reaction condition, green, environmentally friendly, single in product and easy to separate and purify, and the production cost is greatly reduced.

The invention provides a genome editor enDelIscB and a genome editor enDelIscB-T5E based on an IscB system, and further provides a base editor ICBE and a base editor IABE based on the enDelIscBD60A. According to the invention, a genome editor and a base editor which are more compact in size and higher in cleavage activity and insertion / deletion efficiency are realized by optimizing a single mutation site of the activity of a DelIscB protein cleavage plasmid, an optimized variant of the DelIscB protein and an optimized sgRNA variant of the DelIscB. According to the present invention, it is verified that the albinism model is efficiently constructed by using the enDelIscB editor and the enDelIscB-T5E editor through micro-injection of the mouse embryo, such that the enDelIscB editor and the enDelIscB-T5E editor have great potential in the field of genome editing.

The invention discloses a dsPETase01 plastic degrading enzyme mutant and application thereof, and relates to the technical field of bioengineering, the amino acid sequence of a wild type plastic degrading enzyme (dsPETase01) is designed and modified, 8 dsPETase01 single mutation sites or 19 combined mutation sites are provided, the thermal stability of the obtained dsPETase01 mutant is improved by about 4-21 DEG C compared with that of the wild type dsPETase01, and the dsPETase01 mutant can be used for preparing a plastic degrading enzyme. And the activity is improved by nearly 1.2-3 times. The invention also provides four preferable mutant proteins which have higher yield, activity and thermal stability compared with wild dsPETase01, have wider application conditions, are more suitable for degrading PET (Polyethylene Terephthalate) plastics, and are beneficial to large-scale production and industrial application.

The invention discloses construction and application of a novel nitrilase with improved amide generation capability and a mutant thereof. The amino acid sequence of the nitrilase disclosed by the invention is as shown in SEQ ID NO: 1, and the mutant of the nitrilase is obtained by performing single mutation on the 51st site, the 162nd site and the 185th site of the sequence as shown in SEQ ID NO: 1. The nitrilase provided by the invention has excellent amide generation capability, and has important industrial application potential for green and efficient preparation of amide compounds.

The invention discloses a glucose dehydrogenase GOX-2015 mutant and application thereof, and relates to the technical field of bioengineering, the amino acid sequence of wild type glucose dehydrogenase GOX-2015 is designed and modified, that is, 18 single mutation sites and 16 combined mutation sites are provided, and the glucose dehydrogenase GOX-2015 mutant is obtained. Compared with the wild type GOX-2015, the thermal stability of the obtained GOX-2015 mutant is improved by about 2 to 13 DEG C, and the activity of the obtained GOX-2015 mutant is improved by about 1.2 to 4.5 times. The invention further provides a preferable mutant protein which has higher yield, activity and thermal stability compared with wild type GOX-2015, has wider application conditions, is more suitable for efficient production of NADPH coenzyme by a biological conversion method, and is beneficial to large-scale production and industrial application.

The present invention relates to the technical field of enzyme engineering. Disclosed are an artificial intelligence-based enzyme engineering method and system, a device, and a medium. The method comprises: on the basis of structural information of an engineered enzyme to be modified, using an inverse protein folding model to obtain a plurality of structurally compatible protein sequences on the basis of a given scaffold structure; on the basis of the plurality of structurally compatible protein sequences, obtaining a mutant amino acid frequency and a wild-type amino acid frequency at each sequence position; and on the basis of the mutant amino acid frequency and the wild-type amino acid frequency at each sequence position and the structural information of said engineered enzyme, as well as a first creation strategy, a second creation strategy, and a third creation strategy, obtaining a single-mutation variant set and a combinatorial-mutation variant set of said engineered enzyme. The present invention enables efficient and low-cost generation of single-mutation and combinatorial-mutation variant sets of an engineered enzyme to be modified, thereby achieving precise and efficient gene editing and effectively expanding the applicability and safety of gene editing.

The invention discloses a high-temperature-resistant cellobiose epimerase mutant and application thereof. The mutant is obtained by performing single mutation on the 242nd site of an amino acid sequence as shown in SEQ ID No.1. The invention also discloses a preparation method of the high-temperature-resistant cellobiose epimerase mutant. According to the invention, molecular modification is carried out on cellobiose epimerase (DiCE) from Dictyoglomus thermophilum, a mutant with improved thermal stability is screened out, the lactulose conversion efficiency of the obtained mutant is obviously improved, the mutant A242L can tolerate the temperature of 80 DEG C, the lactulose conversion rate is 64.41%, and the industrial application prospect is good.

The invention discloses an L-threonine aldolase mutant and application of the L-threonine aldolase mutant in synthesis of L-serine. The mutant is obtained by performing single mutation on the 88th amino acid of a wild type L-threonine aldolase amino acid sequence shown as SEQ ID No.2. The invention further discloses a preparation method of the L-threonine aldolase mutant. The invention provides the L-threonine aldolase mutant with better catalytic performance, aldol condensation reaction of formaldehyde and glycine can be efficiently catalyzed to prepare L-serine under the condition of not adding a tetrahydrofolic acid precursor, the yield of the L-serine is increased to 72.3% from 56.7% before mutation, the yield is remarkably increased, and the L-threonine aldolase mutant has the advantages that the L-threonine aldolase mutant has a good application prospect. Good application prospects are shown in the synthesis of the L-serine.

The invention provides a tyrosine ammonia lyase mutant and application thereof in production of p-coumaric acid. The tyrosine ammonia lyase mutant is obtained by mutating wild type tyrosine ammonia lyase with an amino acid sequence as shown in SEQ ID No.1, and a mutation site is selected from at least one of Q108S, Y340Q and P375H. The catalytic activity center of wild type tyrosine ammonia lyase is analyzed, and site-directed mutagenesis is carried out on five catalytic activity center sites, so that the mutant shows different enzyme catalytic activity when the mutation sites are different, the mutation sites are the same but the mutation targets are different; q108S, Y340Q and P375H single mutations can significantly improve the enzyme catalytic activity of the tyrosine ammonia lyase mutant, and the tyrosine ammonia lyase mutant having any two mutation sites, especially having three mutation sites, also has better enzyme catalytic activity than the wild type, and has great application potential.

The invention discloses an ethanol dehydrogenase mutant and an application of the ethanol dehydrogenase mutant in synthesis of symmetric chiral diol. The ethanol dehydrogenase mutant is obtained by carrying out single mutation or multiple mutations on amino acids at 74th, 187th, 210th, 211th and 352th sites of an ethanol dehydrogenase amino acid sequence as shown in SEQ ID NO.2 from Candida orbiculata Co. The invention further discloses a preparation method of the ethanol dehydrogenase mutant, and the preparation method comprises the following steps: carrying out single mutation or multiple mutations on the amino acids at the 74th, 187th, 210th, 211th and 352th sites of the ethanol dehydrogenase amino acid sequence as shown in SEQ ID NO.2. Alcohol dehydrogenase is utilized to catalyze prochiral symmetric diketone to synthesize symmetric chiral diol, the method is high in substrate concentration, high in stereoselectivity, mild in reaction, simple in operation process, environment-friendly and more suitable for industrial amplification production, and the problems that in the prior art, the substrate concentration is not high, the reaction condition is violent, the reaction process is complex and difficult to control, and the production cost is low are solved. And the method is not environment-friendly enough, and the like.

The invention discloses a lipase mutant, an engineering bacterium and application of the lipase mutant in preparation of (2S, 3R)-p-methylsulfonylphenyl serine ethyl ester. The lipase mutant is obtained by performing single mutation or combined mutation on 67th, 115th or 400th amino acid of an amino acid sequence as shown in SEQ ID No.1. The invention further discloses a preparation method of the lipase mutant, the engineering bacterium and the application of the lipase mutant in preparation of (2S, 3R)-p-methylsulfonylphenyl serine ethyl ester. The lipase mutant provided by the invention has the advantages that the enzyme activity is obviously improved, the selectivity is not reduced, the reaction conversion rate of the lipase mutant serving as a catalyst reaches 50%, the required reaction time is obviously shortened by 1-7 hours, and the lipase mutant has more excellent catalytic activity. Under the optimal system, the conversion rate after 1000 mM of a substrate is split for 4 h reaches 50.5%, e.e.gt; therefore, a certain basis is provided for industrial preparation of optically pure (2S, 3R)-p-methylsulfonylphenyl serine ethyl ester.

The invention provides a glycosidase mutant and application thereof, the amino acid sequence of the mutant is compared with the amino acid sequence SEQ ID NO.1 of a wild enzyme, in the amino acid sequence SEQ ID NO.1, five sites of M102K, P171L, V203G, T269Y and K299D are respectively subjected to single mutation, pairwise combined mutation, three combined mutation, four combined mutation or one of five combined mutation; the novel glycosidase mutant industrial enzyme is used for synthesis and preparation of hesperetin. The glycosidase mutant enzyme constructed by the invention has the characteristics of low enzyme cost, short conversion time, simple process operation and the like, and has a wide prospect of large-scale industrial application.

The invention relates to the technical field of gene engineering and protein modification, in particular to a high-specific-activity cellulase mutant and application thereof. Compared with the wild type cellulase, the specific activity of the mutants respectively containing single mutation sites of P14A, P19T, S75A, S109N, H118P, M123I, M123L, I130L, I130Q, Y167E, Y167F, Y167W, A176S, A176F, D177S and L192I, provided by the invention, is generally improved by 10.4%-76.1% under a neutral condition. Wherein the specific activity of a single-site mutant containing I130L is the highest, and reaches 183.64 U / mg. The production cost of the cellulase mutant is remarkably reduced, so that wide application of the cellulase mutant is promoted.

Provided are high-specific-activity cellulase mutants and use thereof. Compared to the wild-type cellulase, the specific activity of the provided mutants that comprise single mutation sites P14A, P19T, S75A, S109N, H118P, M123I, M123L, I130L, I130Q, Y167E, Y167F, Y167W, A176S, A176F, D177S, and L192I, respectively, under neutral conditions is generally increased by 10.4% to 76.1%. The single-point mutant comprising I130L has the highest specific activity, which reaches 183.64 U / mg. The production costs of the cellulase mutants are significantly lower, which facilitates the wide use thereof.

The present application relates to the technical field of bioengineering, and particularly relates to a polyphosphate kinase mutant with high catalytic activity and application thereof in synthesis of glutathione. The present application provides a polyphosphate kinase mutant with high catalytic activity, which is characterized in that the polyphosphate kinase mutant is mutated from an amino acid sequence shown in SEQ ID NO: 2 into one of the following: (1) single mutation is made to the 234th amino acid in the amino acid sequence shown in SEQ ID NO: 2; (2) double mutations are made to the 234th and 166th amino acids in the amino acid sequence shown in SEQ ID NO: 2; wherein a nucleotide sequence of a coding gene coding the amino acid sequence shown in SEQ ID NO: 2 is as shown in SEQ ID NO: 1. ch PPK-N234A and double-site mutation polyphosphate kinase mutant ch PPK-N234A / K166D has high activity, and the enzyme activity is increased by 4.19 times and 6.12 times, respectively, compared with wild-type polyphosphate kinase.

The present invention discloses an RthB protein mutant and its application in isoleucine production. The RthB protein mutant is obtained by performing single-site mutations or multi-site mutations at the 16th, 21st, 117th, and 156th positions of the amino acid sequence shown in SEQ ID NO:1. By mutating and modifying the RthB protein, the L-isoleucine fermentation yield of recombinant microorganisms expressing the mutant protein is increased, and the threonine content in the fermentation broth is decreased, thereby improving the yield and purity of isoleucine.

The present invention discloses an L-aspartate aminotransferase mutant and its application. The L-aspartate aminotransferase mutant is obtained by subjecting the amino acid at position 104 of the amino acid sequence of L-aspartate aminotransferase to a single mutation. The L-aspartate aminotransferase mutant is used to synthesize L-phenylglycine derivatives. The L-aspartate aminotransferase mutant is derived from Escherichia coli. After homologous sequence analysis and molecular docking simulation, combined with site-directed mutagenesis verification, a single-point mutation is performed on the amino acid at position 104 to obtain a T104S mutant. The L-aspartate aminotransferase mutant of the present invention can efficiently catalyze the production of L-phenylglycine derivatives. Compared with the wild type, the specific activity of the T104S mutant towards o-chlorophenoxyacetic acid is 3.2 times that of the wild type.

The present invention relates to the field of Biotechnology, particularly to a method based on the introduction of a single mutation in the genes encoding the human IL-2 and the muteins derived thereof that results in increased secretion levels in different hosts without affecting their biological functions. In particular, these mutations are based on a non-conservative change in the amino acid located in position 35 in the primary sequence of human IL-2, preferably the substitutions are K35E, K35D and K35Q. Another object of the present invention are the expression systems used to obtain both the recombinant human IL-2 and the muteins derived thereof using the method described in this invention. The above-mentioned method is useful to improve the production efficiency of the recombinant human IL-2 and the muteins derived thereof both at laboratory and industrial scales. The proteins obtained using this method can be used for therapeutic purposes as well as in the in vitro expansion of T cells for adoptive transfer therapies.

The present invention relates to the technical field of genetic engineering and protein engineering modification, and specifically provides a thermostable acidic pectinase mutant. The acidic pectinase mutant contains a single mutation site S98E. After being treated at 80 °C for 3 minutes, the residual enzyme activity rate is increased by 54.48% compared with the wild type, achieving unexpected technical effects. The thermostability of the acidic pectinase mutant provided by the present invention is significantly enhanced and can be widely applied in fields such as feed and food processing.

The application provides a tyrosine deaminase mutant and application thereof in production of p-coumaric acid. The tyrosine deaminase mutant is obtained by mutating a wild-type tyrosine deaminase with an amino acid sequence shown in SEQ ID No. 1, and the mutation site is selected from at least one of Q108S, Y340Q and P375H. The application analyzes a catalytic active center of the wild-type tyrosine deaminase, and finds that different mutation sites, same mutation sites but different mutation targets can make the mutant show completely different enzyme catalytic activities, and the single mutation of Q108S, Y340Q and P375H can significantly improve the enzyme catalytic activity of the tyrosine deaminase mutant, and the tyrosine deaminase mutant with any two mutation sites, especially with three mutation sites also has excellent enzyme catalytic activity than the wild type, and has great application potential.

The present invention relates to the technical field of genetic engineering and protein engineering modification, and specifically provides an acidic pectinase mutant and its application. The acidic pectinase mutant contains a single mutation site Y219W. After being treated at 80 °C for 3 minutes, the residual enzyme activity rate is increased by 46.77% compared with the wild type, achieving an unexpected technical effect. The heat resistance of the acidic pectinase mutant provided by the present invention is significantly enhanced and can be widely applied in the fields of feed, food processing, etc.

According to the invention, the correlation between Ndufa10 gene and / or S1pr4 gene mutation and muscular dystrophy morbidity is discovered. The invention provides an application of mutations of a Ndufa10 gene and an S1pr4 gene in diagnosis and prediction of myopathy. The invention further provides a muscular dystrophy animal model induced by single mutation and double mutation of the NDufa10 gene and the S1pr4 gene and a modeling method. The invention has application prospects in muscular dystrophy disease diagnosis and therapeutic drug screening.

This invention discloses a tyrosine phenol lyase mutant and its application in synthesizing tyrosine derivatives. The mutant is obtained by single or multiple mutations at position 18 or 417 of the amino acid sequence shown in SEQ ID NO.2. Compared with the wild type, the tyrosine phenol lyase mutant provided by this invention exhibits superior enzyme activity and catalytic performance. The TPL mutant synthesizes levodopa at a cumulative concentration of over 160 g / L, L-tyrosine at a cumulative concentration of over 80 g / L, 3,4,5-TOPA at a cumulative concentration of over 16.4 g / L, 3-fluoro-L-tyrosine at a cumulative concentration of over 33.1 g / L, and 2-fluoro-L-tyrosine at a cumulative concentration of over 20.5 g / L, with an optical purity greater than 99.9%.

The application discloses a 7beta-hydroxysteroid dehydrogenase mutant and application thereof, relates to the field of bioengineering technology, and is designed and reformed according to the amino acid sequence of wild-type 7beta-hydroxysteroid dehydrogenase derived from Collinsella aerofaciens, that is, 17 single-mutation sites or 7 combined-mutation sites of 7beta-hydroxysteroid dehydrogenase are provided, the obtained 7beta-hydroxysteroid dehydrogenase mutant has the thermal stability increased by 2-21.5 DEG C and the activity increased by nearly 1.2-2.24 times compared with the wild-type 7beta-hydroxysteroid dehydrogenase. According to the mutation sites with obvious increase in thermal stability and activity, three preferred mutant proteins are further provided, the three preferred mutant proteins have higher yield, activity and thermal stability compared with the wild-type 7beta-hydroxysteroid dehydrogenase, have wider application conditions, are more suitable for the efficient production of ursodeoxycholic acid by the bioconversion method, and are favorable for large-scale production and industrial application.