54 results about "Sulfotransferase" patented technology

The invention belongs to the technical field of chondroitin sulfate biosynthesis, and discloses a recombinant strain for high-yield chondroitin sulfate, application of the recombinant strain and a preparation method of the chondroitin sulfate. The recombinant strain provided by the invention can be used for simultaneously expressing UDP-glucosamine isomerase, UDP-glucose dehydrogenase, chondroitin polymerase, glucosamine synthetase, phosphoglucosamine mutase, UDP-N-acetylglucosamine pyrophosphorylase, glucose-6-phosphate isomerase and glucose phosphate mutase; according to the present invention, glucose-1-phosphate uridyl transferase, ATP sulfonase, sulfotransferase, pyruvate kinase and APS kinase are adopted, and the enzyme molecules are matched with the original carbon metabolism pathway in the recombinant strain cell to jointly form the enzyme-linked pathway having the efficient CSA synthesis ability, such that the high-yield and high-purity preparation of the CSA can be achieved; the method has the advantages of simple operation, high biological safety of the obtained product, easy realization of large-scale production and the like, and has a good application prospect.

The invention provides recombinant escherichia coli for producing chondroitin sulfate A and application of the recombinant escherichia coli, and relates to the technical field of genetic engineering. According to the recombinant escherichia coli disclosed by the invention, chondroitin synthase, chondroitin 4-O-sulfotransferase, N-acetyl glucosamine epimerase, UDP-glucose dehydrogenase and chondroitin sulfate degrading enzyme are overexpressed; wherein the amino acid sequence of the chondroitin sulfate degrading enzyme is as shown in SEQ ID NO. 5. The recombinant escherichia coli is inoculated into a culture medium for fermentation culture, the average molecular weight of the obtained chondroitin sulfate A is 800Da, and the yield is about 12.5 g / L. Therefore, low-molecular-weight chondroitin sulfate A can be obtained and the yield of chondroitin sulfate A can be increased by fermenting the recombinant escherichia coli disclosed by the invention.

The invention discloses a construction method of a heparin 2-O-sulfotransferase high-activity mutant, and belongs to the technical field of biology. According to the heparin 2-O-sulfotransferase and the application of the heparin 2-O-sulfotransferase, a dissolution promoting label is fused at the N end of the heparin 2-O-sulfotransferase, the coding sequence of the dissolution promoting label is optimized, and active expression of the heparin 2-O-sulfotransferase is achieved; according to the invention, the heparin 2-O-sulfotransferase is mutated, so that a mutant with high stability and catalytic activity is obtained; furthermore, fermentation conditions of the heparin 2-O-sulfotransferase are optimized, so that the expression quantity and the enzyme activity are improved, and the application potential of the heparin 2-O-sulfotransferase in industrial production is enhanced.

The invention relates to the field of biosynthesis of glycosaminoglycan compounds, and discloses a 2-O-sulfotransferase mutant and application of the 2-O-sulfotransferase mutant in heparin synthesis. On the basis of a chicken source 2-O-sulfotransferase protein sequence, site-directed mutagenesis design and modification are carried out, mutation sites comprise 6I, V60I, R120K, G118H, S157H, L227P, E270D, V258I and A275P, and the 2-O-sulfotransferase mutant is obtained. Compared with a wild type, the 2-O-sulfotransferase mutant provided by the invention has the advantages that the expression level, the thermal stability and the catalytic efficiency are obviously improved, and the 2-O-sulfotransferase mutant shows higher conversion rate and structural modification consistency in a reaction system taking N-sulfoheparin precursor polysaccharide as a substrate. The mutant has a wide application prospect in the structural precise control and industrial synthesis process of biological heparin.

The invention discloses a recombinant chondroitin 6-O-sulfotransferase truncation and application thereof in synthesis of chondroitin sulfate C, and belongs to the technical field of bioengineering. On the basis of chondroitin 6-O-sulfotransferase from Norway rats, a chondroitin 6-O-sulfotransferase truncation body is obtained by truncating first 11-106 amino acids at the N end of RnCHST3, the sulfonation rate higher than that of the original RnCHST3 is shown, the sulfonation rate is increased by 51%, and the sulfonation rate in 24 h can reach 11.5%. On the basis of the delta 106RnCHST3, the pET28a (+)-MBP delta 106RnCHST3 for MBP fusion expression is constructed, and the soluble expression quantity of the pET28a (+)-MBP delta 106RnCHST3 is remarkably improved. According to the invention, the chondroitin 6-O-sulfotransferase truncation is used for catalytic preparation of CSC, and a novel green and environment-friendly method is provided.

The invention provides an N-deacetylase / N-sulfotransferase and an application of the N-deacetylase / N-sulfotransferase. According to the invention, systematic stability design and surface electrostatic optimization are carried out on the N-deacetylation structural domain of NDST, and multiple engineering strategies such as structural domain fusion and linker peptide optimization are combined to synergistically improve the specific activity, substrate affinity and bifunctional catalytic synergy of enzyme. Experimental results show that the specific enzyme activity of the modified NDST mutant is obviously higher than that of natural enzyme while the difunctional catalytic capability is maintained.

The invention discloses a construction method of a high-activity mutant of chondroitin 6-O sulfotransferase, and belongs to the field of biological enzyme engineering and synthetic biology. Interaction sites of MusC6 protein and a substrate PAPS are accurately analyzed through a computer-aided molecular docking technology, and key active sites are directionally modified by adopting a strategy of combining rational design and saturated mutation. Experimental results show that the catalytic performance of the obtained mutant S139T / S140L / Y364M / K412V is remarkably improved, and the enzyme activity of the mutant reaches 1182U / L and is improved by 4.4 times compared with the enzyme activity of a wild type 270U / L. According to the invention, the technical bottleneck of low catalytic efficiency of a wild type enzyme on a natural substrate is solved through a multi-site synergistic mutation strategy, and a biocatalyst with industrial application value is provided for efficiently synthesizing chondroitin sulfate C by a microbiological method.

This invention discloses a recombinant 2-O-sulfonyltransferase mutant and its preparation method, belonging to the field of biotechnology. The recombinant enzyme sequence is formed by fusing a truncated (E28~N356) mutant of hamster-derived 2-O-sulfonyltransferase (L119V-R188E-A240I) with an N-terminal maltose-binding protein (MBP) tag. The recombinant 2-O-sulfonyltransferase mutant exhibits higher activity, 1.9 times that of the recombinant wild type, and greater stability, retaining 86.6% of its activity after 5 days at room temperature. Furthermore, the soluble expression level of this recombinant mutant 2-O-sulfonyltransferase is increased by 5 times. This mutant, as a novel biocatalyst with high efficiency, stability, and controllable production costs, has significant application value and broad industrialization prospects in the industrial enzymatic synthesis of glycosaminoglycans such as heparin and heparan sulfate.

The present invention belongs to the technical field of chondroitin sulfate biosynthesis, and discloses a recombinant strain of high-yield chondroitin sulfate and its application and a method for preparing chondroitin sulfate. The recombinant strain provided by the present invention can simultaneously express UDP-glucosamine isomerase, UDP-glucose dehydrogenase, chondroitin polymerase, glucosamine synthetase, phosphoglucosamine mutase, UDP-N-acetylglucosamine pyrophosphorylase, glucose-6-phosphate isomerase, glucose phosphomutase, glucose-1-phosphate uridyl transferase, ATP sulfonase, sulfotransferase, pyruvate kinase and APS kinase, and the above enzyme molecules are combined with the original carbon metabolism pathway in the recombinant strain cell to form an enzyme cascade path with efficient CSA synthesis ability, which can achieve high yield and high purity preparation of CSA, and has the advantages of simple operation, high biological safety of the obtained product and easy large-scale production, and good application prospects.

The application discloses a construction method of a heparin 2-O-sulfotransferase high-activity mutant, and belongs to the technical field of biology.1、The application fuses a solubility-promoting label to the N terminal of heparin 2-O-sulfotransferase, optimizes the coding sequence of the solubility-promoting label, and realizes the active expression of heparin 2-O-sulfotransferase. The application also mutates heparin 2-O-sulfotransferase, and obtains a mutant with high stability and catalytic activity. Further, the fermentation conditions of heparin 2-O-sulfotransferase are optimized, the expression amount and enzyme activity are improved, and the application potential in industrial production is enhanced.

The invention relates to a heparin 3-O-sulfuric acid transferase mutant with high enzyme activity and high thermal stability as well as a coding gene and application of the heparin 3-O-sulfuric acid transferase mutant. The amino acid sequence of the 3-O-sulfate transferase mutant is as shown in SEQ ID NO. 2, and the nucleotide sequence of the coding gene is as shown in SEQ ID NO. 1. The 3-O-sulfuric acid group transferase mutant has remarkable and superior performance in the enzymatic synthesis of heparin glycosaminoglycans. Performance determination shows that the 3-O-sulfate transferase mutant SM1 realizes the synergistic improvement of the catalytic activity and the thermal stability. The total enzyme activity of the 3-O-sulfotransferase is improved by 1.5 times compared with that of wild chicken source 3-O-sulfotransferase. More prominently, the thermal stability of the 3-O-sulfo-transferase is fundamentally improved: the activity of the wild chicken source 3-O-sulfo-transferase is generally and rapidly attenuated within 24 hours at 37 DEG C, while the 3-O-sulfo-transferase mutant SM1 can still retain about 50% of initial activity after being subjected to heat preservation for 8 days under the same condition, so that the 3-O-sulfo-transferase mutant SM1 shows excellent operation stability.

The invention belongs to the field of genetic engineering and synthetic biology, and particularly relates to tyrosine sulfation modified high-activity recombinant hirudin, saccharomyces cerevisiae engineering bacteria and application. According to the invention, saccharomyces cerevisiae is taken as a chassis cell, tyrosine protein sulfotransferase and related enzymes and transport systems are introduced through genetic engineering modification, efficient sulfation modification of the hirudin Tyr63 site is realized, and the anticoagulant activity of the hirudin Tyr63 site is remarkably improved. The method overcomes the limitations of high extraction cost of natural hirudin and lack of modification function in the existing recombination technology. In addition, the tyrosine sulfation modified high-activity recombinant hirudin provided by the invention shows an excellent antioxidant effect in the field of beauty and skin care, and experiments prove that the antioxidant activity of the tyrosine sulfation modified high-activity recombinant hirudin is equivalent to that of natural hirudin. The invention provides an innovative solution for low-cost and large-scale production of high-activity hirudin, and has the application potentials of medicines and cosmetics.

The present invention relates to a method for producing a microorganism having N-deacetylation activity and N-sulfation activity, the method comprising a step (I) of modifying the nucleotide sequence of DNA encoding an N-deacetylase / N-sulfotransferase, and a step (II) of introducing DNA containing the nucleotide sequence modified in step (I) into the microorganism in an expressible manner, wherein the step (I) is a step of codon optimization according to the frequency of use of codons of a biological species different from the microorganism. Furthermore, the present invention relates to a method for producing a microorganism having N-deacetylation activity and N-sulfation activity, the method comprising the step (i) of expressing a specific protein in the microorganism.

To provide a novel inflammasome inhibitor.SOLUTION: An inflammasome inhibitor comprising, as an active ingredient, an inhibitor of 3-mercaptopyruvate sulfurtransferase.SELECTED DRAWING: Figure 2

The invention belongs to the field of genetic engineering and synthetic biology, and particularly relates to tyrosine sulfation modified high-activity recombinant hirudin, saccharomyces cerevisiae engineering bacteria and application. According to the invention, saccharomyces cerevisiae is taken as a chassis cell, tyrosine protein sulfotransferase and related enzymes and transport systems are introduced through genetic engineering modification, efficient sulfation modification of the hirudin Tyr63 site is realized, and the anticoagulant activity of the hirudin Tyr63 site is remarkably improved. The method overcomes the limitations of high extraction cost of natural hirudin and lack of modification function in the existing recombination technology. In addition, the tyrosine sulfation modified high-activity recombinant hirudin provided by the invention shows an excellent antioxidant effect in the field of beauty and skin care, and experiments prove that the antioxidant activity of the tyrosine sulfation modified high-activity recombinant hirudin is equivalent to that of natural hirudin. The invention provides an innovative solution for low-cost and large-scale production of high-activity hirudin, and has the application potentials of medicines and cosmetics.

Compositions and methods are disclosed herein for inducing (up-regulating) the expression of sulfotransferases in the hair follicles, e.g., the scalp. Increasing sulfotransferase is beneficial for metabolizing pro-drugs that require sulfonation to be activated, e.g., minoxidil sulfate is the active metabolite of minoxidil. A method for combining the compositions described herein with topical minoxidil to enhance minoxidil treatment for androgenetic alopecia is described. Additional methods and compositions include the use of retinoid X receptor agonists, retinoic acid receptor agonists, and nuclear receptor agonists in an RXR-NR heterodimer. Additional methods and compositions include the use of a topical solution containing an alkalinizing agent or an alkalinizing agent used with a penetration enhancer for up-regulating the sulfonating capacity of hair bearing skin, hair follicles, and / or keratinocyte cells. In addition, compositions and methods for increasing or decreasing the growth rate of hair follicles is disclosed by altering the intracellular pH. Additionally, methods for preparing a stable liposomal containing solution are described.

The application discloses a method for synthesizing chondroitin sulfate with specific molecular weight by metabolic engineering of Pichia pastoris, and belongs to the technical field of metabolic engineering and synthetic biology. The application uses Pichia pastoris GS115 as a host, constructs a chondroitin synthesis pathway by heterologous expression of kfoA, kfoC and tuaD genes, realizes intracellular synthesis of chondroitin, further strengthens the supply of precursors UDP-GlcA and UDP-GalNAc and redirects carbon metabolism, and improves the yield of chondroitin to 564 mg / L, and the yield in a 5-L fermenter reaches 1.15-5.3 g / L. Further expression of chondroitin sulfate sulfotransferase C4ST and an endogenous phosphorylase system realizes synthesis of chondroitin sulfate with a molecular weight of 40-115 kDa. Finally, by integration of an inducible expression chondroitin lyase ABCI, the control of induction time is 1-20 h, and controllable regulation of chondroitin sulfate molecular weight from 500 Da to 115 kDa is realized.

Compositions and methods are disclosed herein for inducing (up-regulating) the expression and / or activity of sulfotransferases in the hair follicles, e.g., the scalp. The compositions include (i) an agent configured to increase the sulfonating ability of hair bearing skin, hair follicles, and / or keratinocyte cells in a person; (ii) an alkalinizing agent; (iii) liposomes encapsulating at least some of the alkalinizing agent; (iv) an antioxidant configured to prevent oxidative degradation of the liposomes; and (v) a buffer system configured to increase dermal penetration of the liposomes, and / or stabilize the liposome solution; and (vi) a solvent; and (vii) minoxidil or a salt thereof. Additional methods and compositions include the use of the topical solution containing an alkalinizing agent or an alkalinizing agent used with a penetration enhancer for up-regulating the sulfonating capacity of hair bearing skin, hair follicles, and / or keratinocyte cells. In addition, compositions and methods for increasing or decreasing the growth rate of hair follicles is disclosed by altering the intracellular pH. Additionally, methods for preparing a stable liposomal containing solution are described.

The invention discloses a green synthesis method of heparinoid, and relates to the technical field of biological medicine, and the green synthesis method comprises the following steps: S1, dissolving microorganism source chondroitin sulfate in a DES solvent; s2, adding the immobilized sulfotransferase, and carrying out a stirring reaction; and S3, filtering the reaction liquid, freezing the filtrate, and drying to obtain heparinoid powder. The whole reaction is carried out under mild conditions, the conversion rate is high, the reaction time is short, the production efficiency is improved, the use of an organic solvent is reduced through the integrated purification technology, the environmental pollution is reduced, meanwhile, the operation steps are simplified, the production cost is reduced, the high purity of heparinoid is ensured, impurity residues are reduced, and the product quality is improved.

The application discloses a molecular marker related to sulfogroup metabolite content in tea trees and application thereof, and relates to the field of molecular biology. The nucleotide sequence of the molecular marker is shown in SEQ ID NO. 2; and a SNP site exists at the 147th base of the nucleotide sequence, which is a C / A mutation. The application finds a gene CsST2A for controlling the content of multiple sulfogroup metabolites through a whole genome association analysis method, the gene encodes a sulfotransferase CsST2A of tea trees, and develops a molecular marker related to the sulfogroup metabolite content in tea trees based on the same. The molecular marker can be used for rapidly screening tea tree varieties with high sulfogroup metabolite content, and is helpful to shorten the breeding period and improve the breeding efficiency.

The invention relates to a recombinant chondroitin-6-O-sulfotransferase mutant and application of the recombinant chondroitin-6-O-sulfotransferase mutant in synthesis of chondroitin sulfate C, and belongs to the technical field of bioengineering. According to the invention, a dissolution promoting label is fused at the N end of a chondroitin-6-O-sulfotransferase mutant, so that the efficient soluble expression of the enzyme is successfully realized, and the proportion of soluble components is as high as 90%. The mutant HsC6ST-delta loop has the advantages that by carrying out amino acid sequence truncation on chondroitin-6-O-sulfotransferase, the constructed mutant HsC6ST-delta loop has higher chondroitin-6-O-sulfonation activity compared with HsC6ST chondroitin-6-O-sulfonation, the sulfonation rate reaches 78.4% after reaction is carried out for 12 h under the conditions that the temperature is 40 DEG C and the pH is 8.0, and the Tm and half-life period are increased by 9 DEG C and 1.54 h compared with those of an original enzyme.

The invention discloses a benign and malignant thyroid nodule urine marker, an analysis method and application, which can be effectively used for differential diagnosis of benign and malignant thyroid nodules and are noninvasive, high in sensitivity and accurate and reliable in result. The benign and malignant urine marker for thyroid nodules at least comprises one of ubiquitin-like modification activating enzyme 6, sulfotransferase 2B1 and the like.

To obtain a heparin-like substance with high biological activity (anticoagulation activity) as heparin. A method for producing a heparin-like substance is provided, which comprises the step of culturing an animal cell to which the followings are introduced to obtain a culture supernatant containing a heparin-like substance: a polynucleotide encoding glucosaminyl N-deacetylase / N-sulfotransferase (NDST2); a polynucleotide encoding heparan sulfate 3-O-sulfotransferase 1 (Hs3st1); a polynucleotide encoding the extracellular domain of syndecan (SDC); and a polynucleotide encoding 6-O-sulfotransferase.

The invention discloses an MITE transposon for regulating and controlling the content of sulfo metabolite of a tea tree and application of the MITE transposon, and relates to the field of molecular biology. The nucleotide sequence of the MITE transposon is as shown in SEQ ID NO. 1. A whole genome association analysis method finds that insertion variation of an MITE transposon exists in a promoter region of a tea tree sulfotransferase gene. The insertion of the MITE transposon can cause the reduction of the expression quantity of the tea tree sulfotransferase gene, thereby influencing the content of sulfo metabolite. The method for screening the tea tree germplasm with the high sulfo metabolite content is developed based on the insertion variation, the tea tree variety with the high sulfo metabolite content can be rapidly screened, the breeding period is shortened, and the breeding efficiency is improved.

The application discloses a chondroitin-4-O-sulfotransferase mutant construction method and belongs to the field of bioengineering technology. The application is characterized in that a wild-type chondroitin-4-O-sulfotransferase (C4ST) is precisely reformed through a calculation-guided semi-rational design, a SUMO label and an AE connecting peptide are fused at an N end to improve soluble expression, and on this basis, a three-point combined mutant V74L / M117T / Y235V is constructed through a site-directed mutagenesis technology, the enzyme activity of the mutant in a 5L fermenter is as high as 76328 U / L, the specific enzyme activity is 327 U / mg, and the enzyme activity and the specific enzyme activity are 16.6 times and 1.3 times of the wild type, respectively. The mutant is used for catalyzing preparation of chondroitin sulfate A, the sulfation degree can reach 97.0% after 48 h of reaction at 37 DEG C. The application significantly improves the catalytic efficiency of C4ST and provides a key technology of efficient and stable enzyme method for preparing CSA.

The invention discloses a parabacteroides gore DZ001 strain with a cholesterol sulfation function and application of the parabacteroides gore DZ001 strain. The preservation number of the parabacteroides gore DZ001 is GDMCC No: 67376, and the preservation number of the parabacteroides gore DZ001 is H. The strain is separated from faeces of healthy people, a genome carries a sequence for coding sulfotransferase, cholesterol can be specifically catalyzed to generate cholesterol sulfate with higher water solubility, intestinal epithelium reabsorption is blocked, and faeces excretion is promoted. In-vitro experiments show that the strain has excellent cholesterol degradation capacity, gastrointestinal tract tolerance and safe gamma-hemolysis characteristics; in-vivo experiments prove that the traditional Chinese medicine composition can remarkably reduce serum total cholesterol and triglyceride levels of high-fat diet mice, increase high-density lipoprotein cholesterol, improve liver fatty degeneration and regulate blood glucose metabolism. The strain and viable bacteria, supernate or cell-free extract thereof can be widely applied to preparation of cholesterol-reducing health-care foods, medicines and animal feeds, and a new way is provided for prevention and treatment of hypercholesterolemia and related cardiovascular and cerebrovascular diseases.

The application discloses a recombinant chondroitin 6-O-sulfotransferase mutant and application thereof in synthesis of chondroitin sulfate C, and belongs to the technical field of bioengineering. The modification method of the application designs the best mutant through rational protein engineering, modifies the protein structure of the Δ106RnCHST3, overcomes the problem of low natural activity of 6-O-sulfonation of the wild type Δ106RnCHST3, and obtains the best mutant Δ106RnCHST3 F210R / Y310F / A209L / A303S / I173L The sulfonation rate of 24h can reach 46%, which is 3 times higher than that of the wild type, and the industrialization of the biological enzyme preparation CSC is accelerated.

Provided herein are small molecule compounds and methods inhibiting human sulfotransferase 1A3 (SULT1A3) using these small molecule compounds. Methods of manufacturing and treatment are also disclosed.

The present invention relates to a method for producing a microorganism having N-deacetylation activity and N-sulfation activity, comprising the steps of (I) modifying the nucleotide sequence of DNA encoding N-deacetylase / N-sulfotransferase and (II) introducing the DNA containing the modified nucleotide sequence in step (I) into a microorganism so that it can express, wherein step (I) is a step of codon optimization to match the codon usage frequency of a different species of organism than the microorganism. The present invention also relates to a method for producing a microorganism having N-deacetylation activity and N-sulfation activity, comprising the step of (i) expressing a specific protein in the microorganism.

The present disclosure relates to heterocyclic compounds, pharmaceutical compositions comprising said compounds and a method of treating cancer comprising administering the compounds, or compositions thereof to a subject in need thereof. Specifically, the present disclosure provides heterocyclic compounds useful as anti-cancer agents, pharmaceutical compositions comprising said compounds and a method of treating SULT1 positive cancer comprising administering the compounds, or compositions thereof to a subject in need thereof.