10 results about "Glycosynthase" patented technology

The invention belongs to the technical field of genetic engineering and functional enzyme preparations, particularly relates to sucrose synthase and a coding gene and a mutant thereof, and further discloses application of the sucrose synthase. According to the invention, sucrose synthase is obtained from candidate methylobacterium and is further mutated, so that a sucrose synthase mutant of which the enzyme activity, the conversion rate and the stability are remarkably improved is obtained, and the activity and the efficiency of a stevioside synthesis process are effectively improved.

This invention belongs to the fields of genetic engineering and enzyme engineering, specifically relating to a sucrose synthase mutant, its preparation method, product, and application. The mutant is based on the amino acid sequence of the wild-type sucrose synthase shown in SEQ ID NO.1, with the 94th amino acid mutated from P to N, resulting in the amino acid sequence shown in SEQ ID NO.2; or the 660th amino acid mutated from Y to H, resulting in the amino acid sequence shown in SEQ ID NO.3. This mutant can be used for in vitro glycosylation reactions, plant genetic improvement, or industrial biocatalysis, solving the technical problems of poor stability and low catalytic efficiency of natural enzymes.

The present application relates to methods of producing a protein in fucosylated and afucosylated forms at a predetermined ratio by partially or completely inactivating GDP-keto-6-deoxymannose-3,5-epimerase, 4-reductae (GDP-L-fucose synthase, EC 1.1.1.271) and additionally by adjusting the presence or absence of fucose source during culturing.

The invention belongs to the field of gene and enzyme engineering, and particularly relates to an AtSuSy mutant as well as a preparation method, a product and application thereof. The novel sucrose synthase mutant is obtained by mutation of a 782 site of wild sucrose synthase from Arabidopsis thaliana, the amino acid sequence of the wild sucrose synthase is as shown in SEQ ID NO. 1, and the amino acid sequence of the mutant is as shown in SEQ ID NO. 3. Enzymatic property characterization is carried out through plasmid construction, protein expression and purification, the residual activity of the mutant E782P after incubation for 10 min at 65 DEG C is 66.91% which is far higher than 26.51% of wild type sucrose synthase, meanwhile, the relative activity of the mutant E782P is 114.09% of that of wild type sucrose synthase, the activity is improved while the thermal stability is improved, and the mutant E782P has the potential of industrial application.

The invention belongs to the field of gene engineering and enzyme engineering, and particularly relates to a sucrose synthase mutant as well as a preparation method, a product and application thereof. The mutant is characterized in that on the basis of a wild sucrose synthase amino acid sequence as shown in SEQ ID NO.1, the 94th amino acid is mutated from P to N, and the mutated amino acid sequence is as shown in SEQ ID NO.2; or the 660th amino acid is mutated into H from Y, and the mutated amino acid sequence is as shown in SEQ ID NO.3. The mutant can be used for in-vitro glycosylation reaction, plant genetic improvement or industrial biological catalysis, and solves the technical problems of poor stability and low catalytic efficiency of natural enzyme.

The invention discloses a method for preparing rare ginsenoside 20 (S)-Rg3 by directionally converting panax notoginseng stem and leaf saponin based on multienzyme synergistic catalysis, and belongs to the technical field of enzyme engineering. The method comprises the following steps: firstly, generating 20 (S)-Rg3 by using multifunctional xylosidase Tpexy3 and glucosidase Tpebgl3; further, glucosidase Dth3 and glucosidase Tpebgl1 are used for efficiently converting the saponin of the non-Rg3 into the protopanoxadiol; finally, PPD is used as a substrate, and directional synthesis of 20 (S)-Rg3 is achieved through a three-enzyme cascade reaction system composed of UGT mutants MuUGT73F13, UGT94Q2 and sucrose synthase GmSUS. According to the method, the resource advantages of the PPD type saponin in the pseudo-ginseng stems and leaves are fully utilized, the raw material utilization rate and the product added value are remarkably improved, and a feasible path is provided for green and efficient production of the rare ginsenoside.

The invention belongs to the technical field of genetically engineered bacteria, and particularly relates to a multienzyme co-expression genetically engineered bacterium as well as a preparation method and application thereof. According to the invention, genetic engineering bacteria capable of co-expressing sucrose synthase, beta-1, 2-glycosyltransferase and beta-1, 3-glycosyltransferase are constructed, so that fundamental transformation from a traditional complex process of three-bacterium three-enzyme step-by-step fermentation-in-vitro mixing to an innovative mode of one-bacterium three-enzyme one-step catalysis is successfully realized; the innovation of the technical route enables the production efficiency, the cost control and the process simplification to be remarkably improved.

The invention belongs to the field of enzyme engineering, and particularly relates to a thermostable sucrose synthase mutant and application thereof. On the basis of a sucrose synthase amino acid sequence as shown in SEQ ID NO: 1, site-directed mutagenesis is carried out through a rationally designed molecular modification technology, and the thermal stability of the modified sucrose synthase mutant is remarkably improved. The mutant enzyme is obtained by carrying out induced expression and protein purification on the obtained mutant strain, and compared with wild type sucrose synthase, the half-life period of the mutant is prolonged from 42.88 min to 679.41 min under the condition of 50 DEG C and is 15.8 times that of the wild type sucrose synthase; at the temperature of 55 DEG C, the half-life period is prolonged from 12.94 min to 110 min and reaches 8.5 times of that of a wild type, and the strain can better adapt to industrial production.

This invention belongs to the field of enzyme engineering, specifically relating to a thermostable sucrose synthase mutant and its applications. Based on the amino acid sequence of sucrose synthase shown in SEQ ID NO:1, this invention utilizes rationally designed molecular modification techniques to perform site-directed mutagenesis, resulting in a significantly improved thermostability of the modified sucrose synthase mutant. The mutant enzyme was obtained by inducing expression and purifying the protein from the resulting mutant strain. Compared to the wild-type sucrose synthase, the mutant exhibits a half-life that is extended from 42.88 min to 679.41 min at 50°C, which is 15.8 times that of the wild-type; and at 55°C, the half-life is increased from 12.94 min to 110 min, reaching 8.5 times that of the wild-type, thus better suited for industrial production.

The present application relates to glycosyltransferase mutants and their use in preparing quercetin-3-O-rhamnoside, by replacing the 44 amino acids of the C-terminal PSPG domain of the amino acid sequence of the glucosyltransferase AtUGT78D2 with the 44 amino acids of the C-terminal PSPG domain of the glucosyltransferase AtUGT78D1, obtaining a domain-engineered glycosyltransferase AtUGT78D2-D1PSPG; then performing site-directed mutagenesis on the glycosyltransferase AtUGT78D2-D1PSPG, obtaining a rhamnosyltransferase mutant with good selectivity, good stability and high expression, and the present application also provides a three-enzyme coupling catalytic synthesis reaction system for quercetin-3-O-rhamnoside constructed by the UGT78D2-D1PSPG mutant, which realizes the generation of UDP-rhamnose donor from sucrose as a substrate under the action of UDP-rhamnose synthase, reduces the production cost of UDP-rhamnose donor purchase, and then catalyzes the one-step directional synthesis of quercetin-3-O-rhamnoside from quercetin under the action of glycosyltransferase. At SuSy, UDP-rhamnose synthase At RHM-NRS, glycosyltransferase At UGT78D2-D1PSPG mutant The three-enzyme coupling catalytic synthesis reaction system for quercetin-3-O-rhamnoside constructed by the UGT78D2-D1PSPG mutant realizes the generation of UDP-rhamnose donor from sucrose as a substrate under the action of UDP-rhamnose synthase, reduces the production cost of UDP-rhamnose donor purchase, and then catalyzes the one-step directional synthesis of quercetin-3-O-rhamnoside from quercetin under the action of glycosyltransferase.