9results about How to "High substrate concentration" patented technology

The invention belongs to the field of biological pharmacy and biological conversion, and particularly relates to a lipase mutant and application thereof. The lipase CALB mutant disclosed by the invention is obtained by carrying out single-point mutation or combined mutation on the 40th site, the 134 site, the 154 site, the 189th site, the 278th site and the 281 site of an amino acid sequence as shown in SEQ ID NO.1. Compared with a wild type, the mutant has the advantage that the stereoselectivity is greatly improved when the mutant is used for catalyzing racemization 3-cyclohexene-1-methyl formate to produce (S)-3-cyclohexene-1-formic acid. Compared with a traditional chemical method for preparing (S)-3-cyclohexene-1-formic acid, the lipase CALB mutant provided by the invention is high in stereoselectivity, mild in reaction condition, environment-friendly, low in equipment requirement, capable of greatly reducing the production cost and wide in application prospect in industrial application.

The invention relates to a lipase mutant and application thereof in preparation of epoxide, in particular to a lipase mutant which is derived from Tilletia indica and has remarkably improved peroxidation activity, a coding nucleic acid of the lipase mutant, a recombinant expression vector and a recombinant expression transformant containing a nucleic acid sequence, and an application of the lipase mutant in preparation of epoxide. The recombinant lipase or the recombinant expression transformant is used as a catalyst for preparing peroxy acid, and the peroxy acid reacts with olefin to generate epoxide. Compared with other methods (including a chemical method or a biological method and the like) for preparing alpha-epoxy pinane, the enzyme and the technical method thereof disclosed by the invention have the advantages of mild reaction conditions, environmental friendliness, lower cost, simplicity and convenience in operation, easiness in amplification and the like, and have a very good application prospect in the field of producing epoxy products by taking carbon-carbon double bonds as substrates.

The application belongs to the field of biopharmacy and bioconversion, and particularly relates to a lipase mutant and application thereof. The lipase CALB mutant of the application is obtained by adopting single-point mutation or combined mutation on positions 40, 134, 154, 189, 278 and 281 of the amino acid sequence shown in SEQ ID NO. 1. The mutant greatly improves the stereoselectivity in catalyzing the production of (S)-3-cyclohexene-1-methyl formate from racemic 3-cyclohexene-1-methyl formate. Compared with the traditional chemical method for preparing (S)-3-cyclohexene-1-methyl formate, the lipase CALB mutant provided by the application has high stereoselectivity, mild reaction conditions, is friendly to the environment, has reduced requirements on equipment, greatly reduces the production cost, and shows a wide application prospect in industrial application.

The invention belongs to the field of biotechnology and fermentation, and particularly discloses a method for synthesizing ergothioneine through microbial cell membrane ectobiological catalysis and application. According to the invention, L-cysteine and N, N, N-trimethyl histidine are taken as substrates, two catalytic proteins are connected to a microbial cell membrane anchoring protein through a connexon for out-of-membrane expression and tandem catalytic biosynthesis of the ergothioneine, and cell catalytic production of the ergothioneine is successfully realized. Compared with reported organic and biosynthesis methods, the biosynthesis method disclosed by the invention overcomes the requirements that a substrate enters a cell membrane and a product is discharged out of the cell membrane in the traditional intracellular fermentation production, so that the substrate concentration and the product concentration in a cell catalysis process are easy to improve; meanwhile, the interference influence of a substrate and a product on the intracellular environment of microorganisms is reduced, and the strain can be applied to fermentation production, so that the strain is more economical and environment-friendly and has a great market development value.

The invention discloses a method for preparing 5-hydroxymethylfurfural from native wood biomass and purifying the 5-hydroxymethylfurfural, and belongs to the technical field of comprehensive utilization of biomass resources. According to the method, metal salt is adopted as a catalyst, primary woody biomass is subjected to high-substrate-concentration hydrolysis in a single-phase system through a one-pot method, then 5-hydroxymethylfurfural is separated and purified through hydrolysate, and high-yield and high-purity 5-hydroxymethylfurfural is obtained through simple steps such as rotary evaporation, extraction and rectification. According to the method provided by the invention, a large amount of organic extractant is replaced with water, a large amount of humus can be removed, the cost is reduced, the operation is simple, industrialization is easy, and the method is green and environment-friendly.

Provided are a bacillus subtilis genetically engineered bacterium for producing tagatose and a method for preparing tagatose. The genetically engineered bacterium comprises constructing thermostable α-glucan phosphorylases, thermostable glucose phosphomutases, thermostable glucose phosphate isomerases, thermostable 6-tagatose phosphate epimerases, and thermostable 6-tagatose phosphate phosphatases which are independently expressed or coexpressed. The usage of the genetically engineered bacterium can effectively convert starch into tagatose. Compared with existing methods for producing tagatose, the method has advantages such as suitability for whole-cell recycling, high safety, high yield, simple production process, low cost, and easiness in large-scale preparation.

The invention provides transaminase, the amino acid sequence of the transaminase is an amino acid sequence obtained by mutation of an amino acid sequence shown as SEQ ID NO.2, and the mutation at least comprises one of the following mutation sites: the 418th site and the 419th site; the glycine at the 418th site is mutated into methionine, and the threonine at the 419th site is mutated into proline or serine. The transaminase can be used for synthesizing (S)-3-aminobutyronitrile, methyl acetoacetate serves as an initial substrate, (S)-3-aminobutanamide is synthesized through an enzyme method and a chemical method, and finally the (S)-3-aminobutyronitrile and derivatives are prepared through dehydration, the process does not involve extremely toxic substances such as methylsulfonyl chloride, sodium cyanide and the like, post-treatment is simple and efficient, and the method is suitable for industrial production. Finally, a high-purity product is prepared.

The application provides a method for synthesizing dextroketoprofen. The method comprises the following steps: S1: taking ketoprofen amide as a raw material, performing a hydrolysis reaction in a reaction medium, a cosolvent, a phase transfer catalyst and an amidase system, and performing separation and purification to obtain dextroketoprofen and levo-ketoprofen amide; S2: performing a chemical racemization reaction on the levo-ketoprofen amide in the presence of a solvent and a base to obtain ketoprofen amide; and repeating step S1 by taking the ketoprofen amide as a raw material. The method has high substrate concentration, low enzyme substrate ratio, high selectivity, high conversion rate and high product purity, higher efficiency, low solvent consumption, lower cost, fully utilized raw materials, less solid waste discharge, can meet the requirements of industrialized production of enzyme catalysis, and has good industrial application potential.

The application provides an aspartase mutant and a method for producing R-3-aminobutyric acid by using the aspartase mutant, and belongs to the field of enzyme engineering. The amino acid sequence of the mutant is obtained by one or more times of mutation of the sequence shown in SEQ ID NO. 3. The mutant contains one or more of the four mutant sites at positions 237 (T237C), 31 (I31V), 88 (K88N) and 123 (G123S). The aspartase mutant of the application has the advantages of high catalytic activity and high selectivity. The cell obtained by fermentation can catalyze 300 g / L of substrate, which is the highest substrate concentration reported at present. When the aspartase mutant is used for synthesizing R-3-aminobutyric acid, the R-3-aminobutyric acid production rate and the chiral purity can reach more than 99% in 7 hours. Compared with the previous chemical synthesis method and biological enzyme method, the process is simple, the conversion rate is high, the production cost is relatively low, and the aspartase mutant has a good industrial application prospect.