122 results about "Glucosyltransferase" patented technology

The invention discloses a preparation method of glucosyl stevioside, and relates to the technical field of glucosyl stevioside production, rebaudioside A or stevioside is taken as a raw material, cyclodextrin is taken as an auxiliary material, cyclodextrin glucosyltransferase is added for glycosylation reaction, a glucosyl stevioside solution I is cooled, and a glucosyl stevioside solution II is obtained; the method comprises the following steps: adding sucrose, UGT glycosyl transferase, sucrose synthase and uridine diphosphate into rebaudioside A or stevioside, continuously converting unreacted rebaudioside A or stevioside into rebaudioside D or rebaudioside E, inactivating after the reaction is finished, filtering by a plate frame, removing impurities through resin, analyzing the resin by using ethanol to obtain an analysis solution, and treating the analysis solution at 78-82 DEG C for 2-4 hours to obtain the rebaudioside D or rebaudioside E; and dealcoholizing at the vacuum degree of-0.1 MPa, concentrating and drying to obtain the glucosyl stevioside. The quality of the product is improved, the cost is reduced, and the taste of the product is integrally improved.

The invention belongs to the technical field of gene engineering, and particularly relates to a highland barley flavonoid glycosyl transferase gene and application thereof. A novel glycosyl transferase gene HOVUSG6091200 in highland barley is found, and protein expressed by the gene can efficiently catalyze glycosylation modification of various flavonoid compounds, so that flavonoid compound glucoside is prepared, and the health-care value of the highland barley is improved; the gene is also transferred into tobacco, so that the highland barley flavonoid glycosyl transferase gene is also expressed in a tobacco plant, flavonoid glucoside is further generated, and the value of the tobacco plant is improved; the novel gene, the recombinant vector, the recombinant bacteria and the transgenic plant provided by the invention have good application prospects.

The invention discloses a multi-site mutant of glucosyltransferase and an application of the multi-site mutant. A GT 1 family O-glycosyl transferase UGT73C33 derived from liquorice is mutated, an excellent multi-site mutant of the O-glycosyl transferase is obtained through screening, the multi-site mutant contains three mutation sites, F148 is mutated into W, F388 is mutated into W, and A389 is mutated into T. The novel O-glycosyl transferase mutant C33-F148W-F388W-A389T is high in specificity, the selectivity of the novel O-glycosyl transferase mutant C33-F148W-F388W-A389T for generating luteolin-4 '-O-glucoside through specific conversion of luteolin reaches 80%, the novel O-glycosyl transferase mutant is high in catalytic activity, and the yield of luteolin-4'-O-glucoside reaches 156.8 mg / L under suitable conditions. The invention provides a unique biocatalyst and a conversion process thereof for the preparation of luteolin-4 '-O-glucoside with high activity and high added value.

Transgenic plants with blue flower color, or their inbred or outbred progeny, or their propagules, partial plant bodies, tissues or cells, are provided. A buckwheat-derived C-glucosyltransferase (CGT) gene or its homolog is transferred into a host plant to cause delphinidin-type anthocyanins and flavone mono-C-glycosides to be copresent in the plant cells.

The present application belongs to the technical field of oral care, and particularly relates to an oral care composition and a preparation method thereof. The oral care composition comprises, by mass percentage, chitosan oligosaccharide 0.1-0.3%, sea hao polysaccharide 0.08-0.14%, forskolin 0.07-0.12%, corn sprout extract 0.05-0.09%, thickening agent 1-3%, humectant 40-50%, and the balance being water. The oral care composition of the present application can effectively inhibit the activity of glucosyltransferase, inhibit the production of dextran, interfere with the formation process of dental plaque, and reduce the adhesion sites of pathogenic bacteria in the oral cavity. In addition, the composition of the present application can promote the proliferation and repair of oral mucosa epithelial cells, and help repair oral mucosa damage. Furthermore, the use of each component in the composition of the present application can balance the oral microbial flora, while achieving the effect of inhibiting pathogenic bacteria and improving the oral environment.

The invention discloses high-purity resistant dextrin and a preparation method thereof, and the preparation method comprises the following steps: (1) adding 4, 3-alpha-glucosyltransferase into a maltodextrin aqueous solution or a starch solution treated by starch debranching enzyme to react, and after the reaction is finished, deactivating the enzyme; (2) adding alpha-amylase into a reactant in the step (1) to react, and deactivating the enzyme after the reaction is finished; (3) adding alpha-glucosidase into the reactant in the step (2) for reaction, and deactivating the enzyme after the reaction is finished; (4) adding saccharifying enzyme into the reactant in the step (3) to react, and deactivating the enzyme after the reaction is finished; and (5) adding yeast powder into the reactant in the step (4) for fermentation, filtering by a filter membrane after fermentation, collecting membrane filtrate, and drying to obtain the high-purity resistant dextrin. The preparation process is simple and efficient, the product purity is high, the proportion of alpha-1, 3 glucosidic bonds in the prepared resistant dextrin is 40% or above, and the purity of the resistant dextrin is 80-90%.

The invention relates to Escherichia coli for producing 6-alpha-glucosyltransferase and application of the Escherichia coli in production of highly branched starch, and belongs to the technical field of enzyme engineering. According to the invention, a pET20b (+) vector is used for heterologously expressing a 6-alpha-glucosyltransferase coding gene derived from Sporosarcina globispora C11 in escherichia coli, and a signal peptide is designed at the upstream of the gene, so that the secretion and activity of enzyme can be promoted. The Escherichia coli and the microbial agent constructed based on the method can convert high amylose starch into highly branched starch, the reaction condition is mild, and the operation is simple and convenient. By directionally introducing an exogenous gene and optimizing an expression system, the hydrolytic activity of the recombinant 6-alpha-glucosyltransferase reaches 80-100U / mL, the proportion of a starch branched chain structure is remarkably increased, the problems of low modification efficiency and high cost of a traditional enzyme method are solved, and an efficient biological catalysis tool is provided for the fields of food, materials and the like.

Disclosed herein are methods of producing a food product or food precursor. These methods can comprise, for example, (a) providing milk, (b) adding sucrose to the milk to form sweetened milk, (c) contacting the sweetened milk with at least one glucosyltransferase to form insoluble alpha-glucan in the milk, (d) acidifying the milk product to a pH below about 5.0, and (e) inactivating the glucosyltransferase by subjecting the acidified milk product to a temperature of about 70-110 °C. Such methodology can produce an acidified milk product having a stable sucrose level and typically also a stable level of texture. Such methodology can optionally use other types of food products / precursors instead of milk, as appropriate. One or more glucosyltransferases used in the methodology can be selected from (i) a glucosyltransferase that synthesizes alpha-1,6-glucan, and / or (ii) a glucosyltransferase that synthesizes alpha-1,3-glucan. Food products / precursors produced by this methodology are also disclosed.

Pigment compositions comprising betalain pigment molecules are provided. Compositions comprising betanin and betanidin are also provided as are compositions comprising less than 5% sugars by weight. Cells comprising at least one copy of an exogenous tyrosine hydroxylase gene, at least one copy of an exogenous L-DOPA oxidase gene, at least one copy of an exogenous DOPA 4,5-dioxygenase (DOD) gene, at least one copy of an exogenous NADPH cytochrome P450 reductase gene, and at least one copy of a cyclo- DOPA-5-O-glucosyltransferase gene integrated into the genome of the cell are also provided.

A glucosyltransferase catalyzing RA to produce RM and a use thereof are provided. The glucosyltransferases derived from Cornus florida and Juglans regia L. are screened and obtained. It was found that the glucosyltransferase derived from the Cornus florida exhibited a relatively high enzymatic activity in catalyzing the RA to produce the RD, while the glucosyltransferase derived from the Juglans regia L. exhibited a relatively high enzymatic activity in catalyzing the RD to produce the RM. When these two enzymes were used together in catalyzing the RA to produce the RM, a relatively high conversion rate was achieved. The use of the glucosyltransferases derived from the Cornus florida and the Juglans regia L. for preparing the RM of SGs, has not been reported yet. The glucosyltransferase provides an excellent candidate protein resource and its corresponding gene sequence for the enzymatic conversion process of the SGs.

The present invention belongs to the field of biotechnology and specifically relates to mutants of glucosyltransferases and their applications. The mutants of the glucosyltransferases are obtained by combining the amino acid sequence of the glucosyltransferase shown in SEQ ID NO.1 with only one or more of the following mutations: Q60S, Q60A, Q60M, Y452I, Y452V, Y452R, R455K, R455I, R455V, and R455H. Compared to the wild-type, the glucosyltransferase mutants obtained by the present invention have significantly improved catalytic activity and can catalyze maltose to produce maltooligosaccharides. They can be used in the preparation of modified starch to increase the content of slowly digestible starch and resistant starch. Therefore, the present invention has high application value.

The application relates to the field of biological medicine, and discloses a glucosyltransferase mutant, a coding gene of the glucosyltransferase mutant, a recombination carrier, a recombination strain, an enzyme preparation, and application of the glucosyltransferase mutant. The glucosyltransferase mutant is a point mutation at the R position of the amino acid sequence shown in SEQ ID NO:1, and 4-8 histidines are added at the N end. A method for preparing gastrodin comprises the following steps: contacting the glucosyltransferase mutant with p-hydroxybenzyl alcohol and uridine diphosphate glucose. The glucosyltransferase mutant provided by the application has higher specificity for the substrate p-hydroxybenzyl alcohol, thereby obviously improving the conversion rate of the conversion of p-hydroxybenzyl alcohol into gastrodin, and having a good industrial application prospect.

The present invention discloses a genetically engineered bacterium for the whole-cell catalyzed synthesis of glucoside ferulate and its application, belonging to the technical field of bioengineering. In the present invention, by overexpressing the expression of two key enzymes involved in the synthesis of uridine diphosphate glucose (UDP-glucose) in the host cell, namely phosphoglucomutase and uridine diphosphate glucose pyrophosphorylase genes, the intracellular accumulation level of UDP-glucose is further increased; by transferring the UDP-glucosyltransferase gene into the host cell and achieving its high-efficiency expression, a genetically engineered bacterium capable of biocatalytic synthesis of glucoside ferulate is obtained. By adopting the technical scheme of the present invention, the problem of intracellular UDP-glucose supply can be solved, and glucoside ferulate is prepared by whole-cell catalysis using ferulic acid as a substrate. Finally, the yield of glucoside ferulate reaches 220.47 mg / L, and the molar conversion rate is 60.1%.

The invention discloses a saccharomyces cerevisiae genetically engineered bacterium for efficiently synthesizing mogroside V based on multi-compartment multi-metabolic pathways and a construction method of the saccharomyces cerevisiae genetically engineered bacterium. The preparation method comprises the following steps: anchoring an MVA pathway and a squalene synthesis pathway on the surface of a peroxisome membrane, and enabling mogrol synthetase and epoxy squalene cyclization enzyme ERG1 to form a linear polywheat complex through polypeptide interaction; then, through overexpression of MVA pathway key enzyme and introduction of an isopentenol utilization pathway, supply of precursor isopentenyl pyrophosphate and dimethyl allyl pyrophosphate in cytoplasm is increased, and squalene epoxidase ERG1 and mogrol synthetase are anchored in transmembrane ABC efflux protein PDR11 through polypeptide interaction; then anchoring the mogrol synthetase in the lipid droplet subcells; finally, the glucosyltransferase is displayed on the surface of the yeast cell wall through an alpha-lectin system, so that extracellular mogrol is converted into MG-V. The engineering bacterium improves the yield of mogroside V synthesized from the beginning in microorganisms, and has great industrial application value.

The invention provides immobilized cyclodextrin glucosyltransferase as well as a preparation method and application thereof and a preparation method of ascorbic acid glucoside, and belongs to the technical field of enzymes, the technical field of biological catalysis and the technical field of preparation of chemical raw material medicines. The preparation method of the immobilized cyclodextrin glucosyltransferase comprises the following steps: activating a carrier; mixing the activated carrier with a solution containing cyclodextrin glucosyltransferase, stirring, carrying out solid-liquid separation, and washing to obtain an immobilized enzyme; and mixing the obtained immobilized enzyme with a polyethylene glycol-polyethyleneimine copolymer solution, stirring, carrying out solid-liquid separation, and washing to obtain the immobilized cyclodextrin glucosyltransferase. The immobilized cyclodextrin glucosyltransferase prepared by the preparation method is high in catalytic activity and good in use stability, and when the immobilized cyclodextrin glucosyltransferase is applied to catalytic synthesis of ascorbic acid glucoside, high-content and high-yield ascorbic acid glucoside can be obtained.

The present invention relates to a kit for gently detecting 5hmC and a detection method thereof. The kit includes: a blocking reaction solution, an APOBEC deamination conversion solution, a pyridine borane conversion solution, a ligation reaction solution, and an amplification reaction solution; the ligation reaction solution includes probe A and probe B. First, the present invention uses glucosyltransferase to modify uridine diphosphate-glucose (UDP-Glu) on the hydroxyl group of 5hmC to form 5gmc, protecting 5hmC from being affected by subsequent reactions. Then, APOBEC is used to deaminate C and 5mC into uracil (U), and 5fC and 5caC are converted into dihydrouracil (DHU) by pyridine borane. U and DHU are complementary base-paired with A base, while 5gmc is still paired with G, so that 5hmC forms a single-base difference from other bases. The high-sensitivity detection of 5hmC is carried out by loop-mediated isothermal amplification (LAMP) based on ligation. The detection method of the present invention is gentle and efficient, can detect 5hmC as low as 2 fM, and the specificity reaches 1%.

The invention discloses an expression vector of cyclodextrin glucosyltransferase and construction and application of bacillus subtilis, belongs to the technical field of enzyme engineering, and is characterized in that the amino acid sequence of the cyclodextrin glucosyltransferase is shown as SEQ ID NO. 1; the nucleotide sequence for coding the amino acid sequence of the cyclodextrin glucosyltransferase is as shown in SEQ ID NO. 2; the construction method of the expression vector of the cyclodextrin glucosyltransferase comprises the following steps: carrying out whole gene synthesis on an amino acid sequence as shown in SEQ ID NO.1 to obtain a target gene, and connecting the target gene with a vector plasmid pWB980-ori to obtain an expression vector pWB980-ori-CGT; the method has the beneficial effects that the expression vector pWB980-orii-CGT is obtained, the bacillus subtilis is constructed, the cyclodextrin glucosyltransferase can be stably and efficiently expressed, the bacillus subtilis can be directly treated by full-fermentation reaction enzyme liquid without separation, and the constructed genetically engineered bacterium is shortened in fermentation period and has high enzyme activity and high conversion rate.

The present disclosure relates, in part, to addition of glucosyl- or glucano-transferase type enzymes for producing and / or modifying alpha-glucan type structures in dough such as bread dough. Such enzymes encompass, for example, (i) dextrin dextranases, (ii) glucosyltransferases, and (iii) glucosyl-, maltosyl-, maltotetrasyl-, or glucano-transferases, where these enzymes can transfer glucose to / from starch and starch hydrolysate substrates (e.g., dextrin, amylodextrin, and / or maltodextrin) for production and / or modification of alpha-glucan type polymers in bread dough. Further disclosed are methods of preparing a baked food product by baking a farinaceous dough, comprising incorporating into the dough one or more of the foregoing enzymes – e.g., a glucosyl- or glucano-transferase type enzyme – wherein the enzyme can transfer a glucose-based moiety from a polysaccharide or oligosaccharide substrate to the non-reducing end of an oligosaccharide acceptor, thereby improving bread volume and / or dough stability. Further disclosed are modified alpha-glucans for use as food ingredients in general.

The invention provides high-purity hydroxypropyl tetrahydropyrantriol and a synthesis method of the high-purity hydroxypropyl tetrahydropyrantriol. The preparation method comprises the following steps that carbohydrate raw materials and glycosyl transferase are mixed in a buffer solution for an enzymatic reaction, the mass percentage of the carbohydrate transferase in the carbohydrate raw materials is 0.1%-5%, and the carbohydrate transferase comprises one or more of UDP-glucosyltransferase and GDP-mannosyl transferase. And after the enzymatic reaction is finished, adding an organic tin compound such as tributyltin hydride into a reaction system in a protective gas range, and carrying out reduction reaction. And after the reduction reaction, removing most of the solvent in the reaction system to obtain a concentrate. And carrying out supercritical extraction on the concentrate by adopting carbon dioxide as a supercritical fluid and ethanol as an entrainer so as to obtain the high-purity hydroxypropyl tetrahydropyrantriol. The synthesis method can achieve the effects of few reactant residues, no boron residues and low synthesis cost.

The invention discloses an application of UDP (User Datagram Protocol) glucosyltransferase in microbial production of rosavin and rosavin. The UDP glucosyltransferase is used for catalyzing cinnamyl alcohol and UDP-glucose to generate rosai and is coded by an OsUGT13 gene of oryza sativa, and the catalytic efficiency of the UDP glucosyltransferase is 3.6 times that of UDP glucosyltransferase coded by a Bs-YjiC gene from bacillus subtilis. The method can be used for large-scale industrial production of rosavin and the derivative rosavin thereof, and has application value.

The invention discloses a preparation method of glucosyl puerarin, which belongs to the technical field of pharmaceutical chemicals, and comprises the following steps: S1, adding sugar for reaction, mixing puerarin with pure water, adjusting the pH value to 4.5-5.5, adding beta-cyclodextrin, heating to 45-55 DEG C, adding cyclodextrin glucosyltransferase for reaction for 16-22 hours, then adjusting the pH value to 0.1-0.8, adding activated carbon, filtering filtrate and staying overnight at low temperature; s2, desugaring reaction: adjusting the pH value of the filtrate to 3.8-4.5, heating to 55-65 DEG C, adding saccharifying enzyme and glucose oxidase to react for 25-40 minutes, adjusting the pH value to 0.5-1.2, and filtering; s3, column purification: enabling a reaction solution to pass through a mixed column containing sulfonated modified XAD-16 and amino modified HPD-720 resin, and performing graded elution to collect an ethanol eluent; s4, concentrating and drying to obtain a product. The problems that the water solubility of puerarin is poor, the purity of glucosyl puerarin is low, and impurities are difficult to remove can be solved.

The present invention relates to a UDP-glucosyltransferase variant, a fusion enzyme of UDP-glucosyltransferase variants, and a use thereof for producing steviol glycoside. The UDP-glucosyltransferase variant and the fusion enzyme of UDP-glucosyltransferase variants according to the present invention have improved enzymatic activity. In addition, the fusion enzyme of UDP-glucosyltransferase variants has further improved enzymatic activity compared to when each variant is used individually. Accordingly, the UDP-glucosyltransferase variant and the UDP-glucosyltransferase variant fusion enzyme according to the present invention can be widely applied in the field of steviol glycoside synthesis, and have excellent commercial value and significance in that production efficiency can be increased and production costs can be reduced.

The invention discloses a glycosyl transferase mutant for synthesizing rebaudioside M as well as a product and application thereof, and belongs to the technical field of gene engineering. The technical problem to be solved is that the existing glycosyl transferase and sucrose synthase are low in enzyme activity and insufficient in enzyme activity stability, so that the synthesis efficiency of rebaudioside M (RebM) is not high, and the industrial application of preparing RebM by an enzyme method is limited. According to the key points of the technical scheme, the glycosyl transferase mutant is provided, the mutant is obtained by mutating UDP-glycosyl transferase UGT76G1, and the amino acid sequence of the UDP-glycosyl transferase UGT76G is as shown in SEQ ID NO. 1; the mutation comprises mutation on the 83rd site of SEQ ID NO.1, and the amino acid sequence of the glycosyl transferase mutant is shown as SEQ ID NO.5. The invention further discloses a preparation method of the glycosyl transferase mutant.

The invention belongs to the technical field of enzyme engineering, and particularly relates to a cyclodextrin glucosyltransferase mutant and application thereof. The cyclodextrin glucosyltransferase mutant disclosed by the invention is obtained by performing the following mutations on cyclodextrin glucosyltransferase with an amino acid sequence shown as SEQ ID NO.1: aspartic acid at the 251st site is mutated into alanine. The cyclodextrin glucosyltransferase mutant is high in enzyme activity which is 1.2 times that of a wild type, the cyclodextrin glucosyltransferase mutant is applied to hesperidin modification, the substrate conversion rate reaches up to 95.3%, and high-content alpha-glucosyl hesperidin can be obtained after hesperidin modification and enzymolysis of glucosidase.

The present invention provides a cyclodextrin glucosyltransferase mutant, belonging to the technical fields of genetic engineering and enzyme mutants. The mutant is obtained by performing single-point mutation or multi-point mutation on amino acids at positions 197, 345, 407, and 412 of the amino acid sequence shown in SEQ ID No. 1; the mutation includes mutating position 197 of the amino acid sequence shown in SEQ ID No. 1 to H, position 345 to N, position 407 to M, and position 412 to D. The enzyme mutant provided by the present invention can be used to prepare polyglycosylated flavonoids, has a high conversion rate, a simple and convenient purification method, and a low amount of residual glycosyl donors, making it suitable for industrial production and application.

The invention discloses an immobilized recombinant whole-cell catalyst and a method for preparing gamma-cyclodextrin from the immobilized recombinant whole-cell catalyst. The recombinant whole-cell catalyst is prepared by immobilizing recombinant genetically engineered bacterium whole-cells for expressing a high-selectivity gamma-cyclodextrin glucosyltransferase mutant (the amino acid sequence is as shown in SEQ ID NO: 2) in a calcium alginate carrier matrix through an embedding method. The invention also provides a preparation method of the catalyst and application of the catalyst in preparation of gamma-cyclodextrin by catalyzing starch conversion in multi-batch reaction. The immobilized whole-cell catalyst is simple in preparation process and low in cost, has the high enzyme activity utilization rate of free whole cells and the excellent operation stability of immobilized enzyme, the enzyme activity retention rate is still 85% or above after the immobilized whole-cell catalyst is repeatedly used for 8 times at the temperature of 60 DEG C, and the proportion of a gamma-cyclodextrin product can be kept not lower than 96%.

The invention provides glucosyltransferase obtained through a directed evolution method and a derivative, application and a preparation method of the glucosyltransferase, and belongs to the technical field of enzyme engineering. The glucosyltransferase provided by the invention is obtained by a directed evolution method, the amino acid sequence of the glucosyltransferase is a sequence as shown in SEQ ID NO.4, and the DNA sequence of the glucosyltransferase comprises a sequence as shown in SEQ ID NO.3; the glucosyltransferase is high in catalytic capacity, flavone can be remarkably and efficiently catalyzed to generate flavone-7-O-glucoside, and meanwhile generation of precipitates in plant extracts can be remarkably reduced.

The present invention belongs to the field of enzyme engineering technology, and specifically relates to a cyclodextrin glucosyltransferase mutant and its application. The cyclodextrin glucosyltransferase mutant of the present invention is obtained by subjecting the amino acid sequence of the cyclodextrin glucosyltransferase shown in SEQ ID NO.1 to the following mutation: the aspartic acid at position 251 is mutated to alanine. The cyclodextrin glucosyltransferase mutant has a high enzymatic activity, which is 1.2 times that of the wild type. It is applied to the modification of hesperidin, and the substrate conversion rate is as high as 95.3%. After the hesperidin is modified, it can be enzymatically hydrolyzed by glucosidase to obtain a high content of α-glucosyl hesperidin.

The present disclosure relates to a composition for producing glucosylated steviol glycoside, the composition including glucosyltransferase including an amino acid sequence of SEQ ID NO: 1; and a method of producing glucosylated steviol glycoside using the same.

The application provides a cyclodextrin glucosyltransferase mutant and application thereof in preparation of trehalose, belongs to the technical field of genetic engineering and enzyme engineering, and mainly provides a cyclodextrin glucosyltransferase mutant, and amino acid mutation sites are the 33th and 211th amino acids in the cyclodextrin glucosyltransferase amino acid sequence SEQ ID NO.2; the disproportionation characteristic enzyme activity of the cyclodextrin glucosyltransferase mutant provided by the application is obviously improved compared with the wild type, the thermal stability is obviously superior, the optimal temperature of the mutant is 60 DEG C, which is 5 DEG C higher than the optimal temperature 55 DEG C of the wild type, the trehalose yield is obviously improved when the mutant is applied to a multi-enzyme system for preparing trehalose, the substrate utilization rate is improved, and meanwhile, the application prospect of the enzyme in the medical, food and biological industries is improved.