40results about How to "Promotes proper folding" patented technology

The invention relates to a method for recombinating and expressing human fibroblast growth factors, which comprises following steps: fusing an FGF-21 gene and a molecular chaperone Sumo sequence, building a new prokaryotic expression vector, transforming escherichia coli, thereby building engineering bacteria, and obtaining the FGF-21 through culturing the engineering bacteria and inducing expression. The method of the invention can promote soluble expression of protein, which is beneficial for folding recombined protein correctly and is convenient for separating and purifying the protein. Activity detection proves that the recombined protein which is obtained has comparatively high biological activity and the bioactivity is equivalent to the bioactivity of FGF21standard products.

The invention aims to provide a method for improving the expression amount of secretory foreign protein in pichia pastoris. Byintracellular co-expression of double genes of HAC1 and ERO1 or three genes of HAC1, ERO1 and BIP in pichia pastoris, the expression amount of secretory foreign protein in pichia pastoris is effectively improved. According to the invention, double genes of HAC1 and ERO1 or three genes of HAC1, ERO1 and BIP are transformed into pichia pastoris secreting and expressing exogenous xylanase to obtain recombinant pichia pastoris strains, thereby significantly increasing the expression amount of the exogenous xylanase. During co-expression of double genes of HAC1 and ERO1 in pichia pastoris, the secretion expression amount of exogenous xylanase is generally increased by 15%-25% compared with the initial level; and during the co-expression of three genes HAC1, ERO1 and BIP, the secretion expression amount of xylanase is increased by 45%-57% compared with the initial level.

The invention discloses a recombinant plasmid capable of expressing a soluble human papilloma virus 16 subtype L1 protein and an expression method thereof. The recombinant plasmid is constructed by inserting a human papilloma virus 16 subtype L1 gene and an SUMO label gene into an expression plasmid. According to the recombinant plasmid disclosed by the invention, the HPV16 L1 gene is optimized, so that the HPV16 L1 gene is more applicable to efficient expression of a target protein in an escherichia coli host; meanwhile, an Sumo label expression system is initially applied to fusion expression in escherichia coli, and the solubility of the target protein is further enhanced, so that the expressed target protein has higher activity. The expression method disclosed by the invention has the advantages that the target protein solubility expression efficiency is obviously improved, the product property is uniform and stable, the target protein expression quantity in a supernatant of a fermented disrupted bacterium liquid can reach 70Mug / ml, and the numerical value is obviously higher than yields of the target protein in other prokaryotic expression systems, so that the industrial production requirement is met.

The invention relates to a virus-like particle (VLP) of a recombined human papillomavirus (HPV) 33 and a preparation method of the virus-like particle. According to the specific technical key points, a polynucleotide gene segment of a new coded and recombined HPV33 L1 protein, a carrier containing the gene segment, a host cell comprising the carrier, an HPV33 L1 fusion protein translated and expressed by the gene segment, a pentamer and the VLP consisting of the pentamer are provided. The invention furthermore discloses applications of the pentamer, a VLP protein and a vaccine composition consisting of the pentamer and the VLP protein to the preparation of drugs for preventing HPV33 infection.

The invention relates to an expressing method and application of micromolecule thioesterase, in particular to recombinant expression of micromolecule thioesterase to escherichia coli, and applicationof micromolecule thioesterase to production of 10-hydroxy-2-caproleic acid intermediate product trans-2-caproleic acid. Escherichia coli ester acyl coenzyme A thioesterase gene ydiI is connected ontopET-28a plasmid containing a sumo label; the escherichia coli BL21(DE3) is transferred; the recombinant escherichia coli (containing pET-28a-ydiI)) is used; trans-2-caproleic acid is produced througha cell-free system.

The invention discloses a method for preparing recombinant heparinase III by utilizing an SUMO fusion expression system and the heparinase III prepared by the method, the preparation method comprises the following steps: selecting a heparinase III sequence from Pedobacter heparinus, wherein the amino acid sequence of the heparinase III is shown as SEQ ID No.1, and the total number is 659aa; removing a signal peptide sequence to obtain a DNA sequence of the heparinase III, such as SEQ ID No.2; inserting the DNA sequence of the heparinase III into a pSMART vector plasmid with an N-terminal SUMO protein tag; transforming the correct plasmids into BL21 (DE3) escherichia coli competent cells, selecting monoclone, and obtaining fusion protein through fermentation and purification; and cutting the SUMO tag protein from the fusion protein by using the SUMO protease to obtain the heparinase III. The method has the advantages that the solubility of the target protein is improved, correct folding of the target protein is promoted, and inclusion bodies are prevented from being formed; the purification cost is reduced and the purification efficiency is improved; the molecular weight of the SUMO protein tag is small, the influence on the enzymatic activity of the heparinase III is small, and the purified heparinase III is obtained.

The invention belongs to the technical field of genetic engineering, and specifically discloses a genetically engineered bacterium for expressing solubility pig gamma-interferonPoIFN-gamma and construction method and application of the genetically engineered bacterium. The genetically engineered bacterium for expressing solubility pig gamma-interferonPoIFN-gamma is Escherichia coil BL21(DE3) carrying recombinant plasmid Pet-32a(+)-PoIFN-gamma and plasmid pTf16-ParaB-tig simultaneously, wherein the PoIFN-gamma gene sequence in the recombinant plasmid Pet-32a(+)-PoIFN-gamma is SEQ ID NO.4, and the plasmid pTf16-ParaB-tig can express the molecular chaperone tig. The method disclosed by the invention is capable of improving the solubility and output of the pig gamma-interferon, simple in process and low in cost, and has good industrial application value.

The invention relates to a recombinant VLP of HPV31 and a preparation method thereof. Specifically, the invention discloses a novel polynucleotide gene segment coding recombinant HPV31 L1 protein, a vector containing the gene segment, a host cell containing the vector, HPV31 L1 fusion protein translated and expressed by the gene segment, a pentamer and a VLP composed of the pentamer; and the invention also discloses application of the pentamer, VLP protein and a vaccine composition composed of the pentamer and the VLP protein in preparation of drugs used for preventing HPV31 infection.

The invention belongs to the field of medicine and discloses recombinant antibacterial peptide, and a preparation method and an application of the recombinant antibacterial peptide. An amino acid sequence of the recombinant antibacterial peptide is as shown as SEQ ID NO: 1 (Sequence Identifier Number 1). The polypeptide has a better broad-spectrum antibacterial action and can be used for preparing an antibacterial medicine.

The invention discloses construction and an application of an astaxanthin synthesis strain. The invention provides a recombinant bacterium, which is prepared according to a method comprising the following steps of: expressing and synthesizing carotenoid related genes crtY gene, crtZ gene and crtW gene in escherichia coli, and regulating and controlling the expression of molecular chaperone genes groES and groEL in the escherichia coli to obtain the recombinant bacterium, according to the invention, the promoters with different intensities are utilized and gene copy is changed, so that coordinated expression among genes in a synthetic pathway of astaxanthin is realized, and the toxicity of metabolic intermediates is eliminated, so that the generation capacity of biosynthesis of astaxanthin is improved. In addition, stabilizing the expression of the exogenous gene and increasing the correct folding of the exogenous gene can also become an effective way for improving the biological activity of astaxanthin synthetic pathway enzyme and increasing the yield of astaxanthin.

The invention relates to a recombinant human papilloma virus 52 virus-like particle and a preparation method of the recombinant human papilloma virus 52 virus-like particle. The specific technical point is that the invention provides a polynucleotide gene segment of a novel coded recombinant HPV52 L1 protein, a carrier comprising the gene segment, a host cell comprising the carrier, an HPV52 L1 fusion protein translated and expressed by the gene segment, a pentamer and a VLP comprising the pentamer. The invention further discloses application of the pentamer, the VLP protein and a vaccine composition comprising the pentamer and the VLP protein to the preparation of a medicine used for preventing HPV52 infection.

The invention discloses human CKSCFV (creatine kinase single-chain variable fragment) with a molecular chaperone function, which comprises two amino acid sequences. The invention further provides a gene sequence, a carrier and a host cell of the human CKSCFV, and specifically describes the operation process. The human CKSCFV can be interacted with human creatine kinase to facilitate correct folding of the human creatine kinase, and fine effects are achieved. The CKSCFV has fine application prospect and is applicable to the production process of human creatine kinase drug additives.

The invention relates to an 11-type recombinant human papilloma virus virus-like particle and its preparation method. The specific technical key point is to provide a new recombinant coded polynucleotide gene fragment of an HPV11 L1 protein, a vector containing the gene fragment, a host cell containing the vector, an HPV11 L1 fusion protein translated and expressed by the gene fragment, a pentamer and VLP composed of the pentamer. The invention also discloses an application of the pentamer, a VLP protein and a vaccine composition composed of the VLP protein in the preparation of drugs for preventing HPV11 infection.

The invention belongs to the fields of medicine and bioengineering, and discloses an immunoregulation peptide, and a preparation method and application thereof. The amino acid sequence of the immunoregulation is disclosed as SEQ ID NO:1. The immunoregulation peptide has high T lymphopoiesis promoting function, and can be used for preparing T lymphopoiesis promoting drugs.

The invention discloses an escherichia coli soluble expression vector capable of efficiently obtaining recombinant protein. A hexahistidine label-flexible linker peptide-chitin binding domain-peptide-containing complex coding gene and a thioredoxin-hexahistidine label complex coding gene are placed in the same vector, the characteristics that the peptide-containing protein can realize self-cutting under the condition of change of temperature and pH and the thioredoxin can promote correct folding of protein containing more disulfide bonds are used, through affinity mediation of a hexahistidine label or a chitin binding domain, by using an affinity chromatographymethod, the target protein purificationefficiency and purity can be greatly improved, the problem that Escherichia coli gene engineering expression products form inclusion bodies easily is solved at a low cost, the application value is high, and the application range is wide.

The invention discloses a yeast engineering bacteria for producing trypsin in a high-yield manner and a construction method of the yeast engineering bacteria, belonging to the genetic engineering field. According to the constructed yeast engineering bacteria, trypsin of thioredoxin is blended at the front expression end, and four aspartic acids and lysine (D4K) are inserted between the thioredoxin and the trypsin; the trypsin amidase activity (BAPNA (nitroaniline)) is 47.4U / mL and the esterase activity (BAEE (N-benzoyl-L-arginine-ethylester)) is 2667U / mL, the problems of trypsinogen activation and low yield are solved, and the trypsin production method by the yeast engineering bacteria has the advantages of high yield, simplified process and industrial application convenience.

The invention aims to provide a modified duck circovirus Cap protein as well as a preparation method and application thereof, that is, 21-36 peptide fragments rich in arginine at the N-terminal of theduck circovirus Cap protein are deleted, and a universal T cell epitope is introduced into a deletion region to promote immunogenicity; the amino acid sequence of the obtained modified duck circovirus Cap protein is SEQ ID NO: 1, and one nucleotide sequence is SEQ ID NO: 2. The modified duck circovirus Cap protein gene is connected with an expression vector and then transferred into escherichia coli BL21 (DE3), efficient soluble expression of duck circovirus Cap protein is achieved, the expressed protein is spontaneously assembled into virus-like particles, and the particles can be used for development of products such as duck circovirus gene engineering subunit vaccine and yolk antibody.

The invention belongs to the technical fields of genetic engineering, molecular biology and other biology, and particularly relates to an efficient culture medium for expressing a soluble recombinantprotein by escherichia coli and a preparation method of the culture medium. The culture medium can improve the success rate and yield of expressing the soluble recombinant protein in cytoplasm by theescherichia coli; the efficient culture medium is selected from an M9 culture medium, basic ingredients such as sorbitol, ethanol, glucose and sucrose are added, magnesium sulfate and ampicillin are also added, IPTG is used as an inducer, and the efficient culture medium aims to increase the soluble expression of the foreign protein; the efficient culture medium is used to express the recombinantprotein, especially to express the soluble recombinant protein by the escherichia coli; and the culture medium can use escherichia coli BL-21 or DH5alpha strains as reaction bacteria, or K12 or origamiB escherichia coli strains as reaction bacteria.

The invention provides a virus-like particle as well as a preparation method and application thereof, and belongs to the technical field of agricultural science, animal husbandry and veterinary science. Small ubiquitin-like modified proteins are used as tag proteins to promote correct folding of virus structural proteins VP0, VP1 and VP3, meanwhile, the structural stability of the proteins is guaranteed, mass expression of the structural proteins is achieved, and after in-vitro self-assembly of the three structural proteins, a large number of virus-like particles similar to natural viruses in performance are obtained. Immunogenicity detection shows that the prepared virus-like particles have high immunogenicity and can be used as reserve vaccines to be applied to prevention and control of swine vesicular disease or foot-and-mouth disease virus transmission.

The present invention relates to type 33 recombinant human papillomavirus virus-like particles and a preparation method thereof. The specific technical point is to provide a new polynucleotide gene fragment encoding a recombinant HPV33 L1 protein, a vector comprising the gene fragment, and a vector including the vector. Host cells, as well as the HPV33 L1 fusion protein, pentamer and VLP composed of the pentamer translated and expressed by the gene fragment, the present invention also discloses that the pentamer, the VLP protein and the vaccine composition composed of the pentamer are used in the preparation of prevention Drug application in HPV33 infection.

The present invention relates to a method for expressing the HMGB1 A-box protein using the SUMO system, comprising the following steps: using a plasmid containing HMGB1 as a template, designing primers to amplify the A-Box domain, substituting the amplified target gene A-Box Cloned into the pSumo-Mut expression vector between Stu I and Hind III to obtain the pSumo-Mut-A-Box plasmid; use the recombinant plasmid to transform ArcticExpress TM (DE3) Prokaryotic host bacteria, using IPTG to induce expression of the target protein; sumo protease enzymatic hydrolysis to obtain high-purity A-Box protein. The invention has the advantages that: the HMGB1 A-box fusion protein is efficiently, stably and solublely expressed by using the SUMO expression system, and a mature protein with high purity can be obtained after cleavage by SUMO protease I, and no amino acid residue remains.

The invention discloses the construction of an astaxanthin synthetic strain and its application. The invention provides a recombinant bacterium, which is prepared according to a method comprising the following steps: expressing the synthetic carotenoid-related genes crtY gene, crtZ gene and crtW gene in Escherichia coli, and regulating the expression of molecular chaperone genes groES and groEL in the Escherichia coli , the obtained recombinant bacteria; the present invention realizes the coordinated expression of genes in the astaxanthin synthesis pathway by using promoters of different strengths and changing the gene copy, eliminates the toxicity of metabolic intermediates, thereby increasing the production capacity of astaxanthin biosynthesis. In addition, stabilizing the expression of exogenous genes and increasing the correct folding of exogenous genes can also be an effective way to improve the biological activity of enzymes in the astaxanthin synthesis pathway and increase the production of astaxanthin.

The invention relates to a method for recombinating and expressing human fibroblast growth factors, which comprises following steps: fusing an FGF-21 gene and a molecular chaperone Sumo sequence, building a new prokaryotic expression vector, transforming escherichia coli, thereby building engineering bacteria, and obtaining the FGF-21 through culturing the engineering bacteria and inducing expression. The method of the invention can promote soluble expression of protein, which is beneficial for folding recombined protein correctly and is convenient for separating and purifying the protein. Activity detection proves that the recombined protein which is obtained has comparatively high biological activity and the bioactivity is equivalent to the bioactivity of FGF21standard products.

The invention discloses disulfide bond protein based on Gluc luciferase and a preparation method. The method comprises the following steps: introducing SEP-tag to the terminal C of Gluc luciferase gene; inserting the Gluc luciferase gene connected with an SEP tag into a multi-cloning site of an expression vector to obtain a luciferase Gluc-SEP gene expression vector; transforming the expression vector into an escherichia coli strain with trxB<-> / gor<-> resistance gene and DsbC redox gene to obtain an expression strain containing the expression vector; and cultivating the expression strain, and expressing to obtain disulfide bond protein based on Gluc luciferase. The method solves the problems of low optical stability of luciferase and quick fluorescence quenching.

The invention relates to type 45 recombinant human papillomavirus virus-like particles and a preparation method thereof. The specific technical points are to provide a new polynucleotide gene segment encoding a recombinant HPV45 L1 protein, a vector containing the gene segment, and a carrier including the vector. The host cell, and the HPV45 L1 fusion protein, pentamer and VLP composed of the pentamer translated and expressed by the gene fragment, the present invention also discloses the pentamer, VLP protein and the vaccine composition composed thereof in the preparation of the prophylaxis Drug application for HPV45 infection.