25 results about "Pantoic acid" patented technology

The invention discloses a producing strain of D-pantoic acid lactone hydrolase and application of the producing strain. The producing strain is fusarium moniliforme DPLH6, and the preservation number of the producing strain is CGMCC (China General Microbiological Culture Collection Center) No. 42135. The fusarium moniliforme strain disclosed by the invention has the capability of producing the D-pantoic acid lactone hydrolase at high yield, the D-pantoic acid lactone is prepared after the D-pantoic acid lactone hydrolase fermented by the strain is immobilized by glutaraldehyde, less enzyme dosage and shorter catalysis time can be realized, and the ee value of the target product D-pantoic acid lactone is greater than 99%.

The invention provides a method for producing DL-2, 4-dihydroxy-3, 3-dimethyl butyric acid (DL-pantoic acid) and / or DL-3-hydroxy-4, 4-dimethyl oxacyclopentane-2-one (DL-pantolactone), in particular to a method for producing DL-2, 4-dihydroxy-3, 3-dimethyl butyric acid and / or DL-3-hydroxy-4, 4-dimethyl oxacyclopentane-2-one (DL-pantolactone) by condensation of hydroxypivalaldehyde and formyl-CoA of C1 source, in particular to a method for producing DL-2, 4-dihydroxy-3, 3-dimethyl oxacyclopentane-2-one (DL-pantoic acid) and / or DL-3-hydroxy-4, 4-dimethyl oxacyclopentane-2-one (DL-pantoic acid). The invention relates to a method for preparing 2, 4-dimethyl oxacyclopentane-2-one. The present invention also provides a genetically engineered microorganism for use in the production of DL-2, 4-dihydroxy-3, 3-dimethylbutyric acid and / or DL-3-hydroxy-4, 4-dimethyl oxacyclopentane-2-one, and also provides a genetically engineered microorganism for use in the production of DL-3-hydroxy-4, 4-dimethyl oxacyclopentane-2-one. In addition, the key enzyme has been engineered to improve its performance in the production of DL-pantoic acid or DL-pantolactone.

Provided are a genetically engineered bacterium for high-yield production of D-pantothenic acid (D-PA), and a preparation method therefor and a use thereof. The D-PA production yield of the engineered bacterium is significantly improved by means of the following actions: overexpressing acetolactate synthases IlvB and IlvN for pyruvate diversion while deleting a transcription attenuation region and mutating a valine feedback inhibition site; overexpressing ketol-acid reductoisomerase IlvC and dihydroxy acid dehydratase IlvD, and altering the coenzyme preference of IlvC from NADPH to NADH to unlock the upstream pathway for D-PA synthesis; overexpressing PanB and overexpressing a serine-glycine transport system to promote the synthesis of 5,10-methylenetetrahydrofolate; at the same time, enhancing the expression of ketopantoic acid reductase PanE; and finally overexpressing pantothenate synthetase PanC and increasing the content of cofactor ATP required by PanC. By means of the integrated enhancement of the above multiple modules, the provided engineered bacterium achieves improved cell growth due to balanced cofactors and increased D-PA.

The invention discloses a synthesis and preparation method of magnesium tritrip acid, which comprises the following steps: S1, heating magnesium hydroxide and 4-aminobutyric acid in methanol for reaction to obtain magnesium 4-aminobutyrate; s2, reacting the magnesium 4-aminobutyrate obtained in the step S1 with pantoic acid lactone in isopropyl alcohol under heating to obtain a magnesium furonate initial product; and S3, purifying the magnesium furonate initial product to obtain a magnesium furonate finished product. According to the invention, 4-aminobutyric acid, magnesium hydroxide, pantothenic acid lactone, ethanol, methanol and the like are used as raw materials, and the magnesium furonate is prepared through the processes of acid-base neutralization, ester exchange, refined crystal transformation and recrystallization; the method has a series of advantages of good process stability, excellent reaction selectivity, high yield, cheap and easily available raw materials, simple operation, high safety, environmental friendliness and the like.

The invention discloses a pantothenic acid synthetase mutant as well as a preparation method and application thereof, and relates to the technical field of pantothenic acid synthesis. The mutant is obtained by mutating V at the 62nd site of an amino acid sequence as shown in SEQ ID NO.2 into A. By using the pantothenic acid synthetase mutant disclosed by the invention, the enzyme activity of the pantothenic acid synthetase is remarkably improved, and the conversion rate of D-pantoic acid and the yield of D-pantoic acid are improved.

The invention discloses an L-pantoic acid lactone dehydrogenase mutant and an application of the L-pantoic acid lactone dehydrogenase mutant. The L-pantoic acid lactone dehydrogenase mutant is subjected to any one or a combination of at least two of the following mutations on the basis of an amino acid sequence SEQ ID NO.1: T271D, A27S, T163K, V240I or L253I. The L-pantoic acid lactone dehydrogenase mutant is constructed on the basis of the L-pantoic acid lactone dehydrogenase derived from amycolatopsis methyloralis, the mutant shows excellent catalytic activity, the cell apparent activity of the mutant is improved by more than 10 times compared with that of a wild type, and the mutant provides a new biocatalyst for efficient preparation of D-pantoic acid lactone and derivatives thereof.

The present invention introduces a heterologous formate-tetrahydrofolate ligase gene into a microorganism having a D-pantoic acid synthetic metabolic pathway, and assimilates one molecule of formate to generate one molecule of 5,10-methylenetetrahydrofolate, and uses the microorganism to produce a new process of ketopantoate and / or D-pantoic acid from formate and glucose.

The present application relates to the technical field of biological catalysis, and discloses application of L-pantoic acid lactone dehydrogenase and a mutant thereof in preparation of D-pantoic acid lactone, wherein the L-pantoic acid lactone dehydrogenase and the mutant thereof with improved activity are screened, and ketopantoic acid lactone reductase is co-expressed to catalyze production of D-pantoic acid lactone.The present application adopts a method for co-expressing L-pantoic acid lactone dehydrogenase mutants and ketopantoic acid lactone reductase to catalyze synthesis of D-pantoic acid lactone, and has the advantages of simple process, high enzymatic reaction selectivity, no by-product, simple product extraction and refining, and the like.

The invention provides an engineering bacterium for producing D-pantothenic acid as well as a construction method and application of the engineering bacterium. The engineering bacterium expresses an acetolactate synthase large subunit gene ilv2, an acetolactate synthase small subunit gene ilv6, a keto acid reductoisomerase gene ilvC, a dihydroxy acid dehydratase gene ilvD, a hydroxymethyltransferase gene ecm31, a keto pantoic acid reductase gene panE, an L-aspartic acid decarboxylase gene panD and a pantothenic acid synthase gene panC. According to the method, mitochondria is selected as a targeting compartment, and modular combination optimization approach positioning is adopted, so that the yield of D-pantothenic acid is greatly increased, compared with the yield of an original strain without spatial tissue optimization, the yield is increased by more than one time, the metabolic flux bottleneck in a traditional cytoplasm synthesis mode is broken through, and efficient synthesis of D-pantothenic acid is realized.

A 3-methyl-2-oxobutanoate hydroxymethyltransferase variant, a microorganism having enhanced activity of 3-methyl-2-oxobutanoate hydroxymethyltransferase, a composition for producing pantothenic acid and / or pantoic acid comprising the microorganism, and a method for preparing pantothenic acid and / or pantoic acid comprising culturing the microorganism are provided.

The invention belongs to the field of biological medicine, and particularly relates to an engineering bacterium for producing D-pantoic acid lactone hydrolase and a culture and whole-cell immobilization method thereof, the engineering bacterium takes E.coli BL21 (DE3) as a host and pET28a (+) as a carrier, and the D-pantoic acid lactone hydrolase with a nucleotide sequence shown as SEQ ID NO.1 is expressed. The engineering bacterium can efficiently secrete the D-pantoic acid lactone hydrolase, and the engineering bacterium can be used for pilot-scale high-density fermentation culture, so that the industrial production of the D-pantoic acid lactone hydrolase becomes possible. According to the fermentation culture method disclosed by the invention, the supply of a carbon source and a nitrogen source is controlled by screening a fermentation culture medium and controlling the replenishing time, replenishing rate and dosage of glycerol and ammonia water, and the supply of oxygen is controlled by adjusting the stirring rotating speed and ventilation volume, so that the high-density growth of thalli and the efficient expression of target protease are realized. The invention further discloses an immobilization method of the D-pantoic acid lactone hydrolase, and the stability of the D-pantoic acid lactone hydrolase is improved by 32.12%. The production cost is greatly reduced, and the method has practical industrial production value.

The invention discloses a method for recovering D-calcium pantothenate mother liquor. The recovery method comprises the following steps: S1, carrying out pretreatment, precipitation treatment and hydrolysis treatment on D-calcium pantothenate mother liquor to obtain a reaction solution 1; s2, carrying out lactonization reaction on a mixed solution of the reaction solution 1 and a solvent to obtain a reaction solution 2; and S3, carrying out solid-liquid separation on the reaction liquid 2 to obtain a solid, namely beta-alanine, and crystallizing the obtained liquid to obtain the D-pantoic acid lactone. According to the recovery method, high-purity beta-alanine and D-pantoic acid lactone can be obtained, and meanwhile, the recovery rate of the beta-alanine and the D-pantoic acid lactone is effectively increased; in the whole process, a large amount of acid is not needed for neutralization reaction, extraction operation is not needed, the input of organic solvents is reduced, and the production cost is effectively reduced.

Provided are a genetically engineered bacterium with a high yield of D-pantothenic acid based on one-carbon unit supply, a construction method therefor, and the use of said genetically engineered bacterium in the preparation of D-pantothenic acid by microbial fermentation. By enhancing a glycine cleavage system, the supply of a one-carbon cofactor in Escherichia coli organisms is enhanced, so that the carbon flux flows to the synthesis of D-pantothenic acid. In order to continuously enhance the carbon flux, genes related to a THF synthesis pathway are enhanced to regulate a serine synthesis pathway; and by combining the enhancement of the glycine cleavage system and the expression of key genes of a pantoic acid synthesis pathway, a plasmid-free and antibiotic-free engineered bacterial strain used for high-yield production of D-pantothenic acid is finally obtained. Finally, the titer of D-pantothenic acid in shake flask culture is increased by 31.7% relative to that of the original strain and reaches 6.85 g / L, and after fermentation in a 5 L fermentation tank for 92 hours, the D-pantothenic acid yield can reach 112.62 g / L.

The present invention provides a genetically engineered pantoic acid-producing strain having or having an enhanced NADH-dependent acetohydroxy acid reductoisomerase, a method for producing the strain, a method for producing D-pantoic acid using the strain, and use thereof in production of D-pantoic acid.

The present invention provides a genetically engineered pantoic acid-producing strain having or having an enhanced NADH-dependent acetohydroxy acid reductoisomerase, a method for producing the strain, a method for producing D-pantoic acid using the strain, and use thereof in production of D-pantoic acid.

Provided are: a polypeptide having activity of 3-methyl-2-oxobutanoate hydroxymethyltransferase; a microorganism having increased activity of 3-methyl-2-oxobutanoate hydroxymethyltransferase; a composition for producing pantothenic acid and / or pantoic acid, comprising the polypeptide and / or the microorganism, and a pantothenic acid and / or pantoic acid production method comprising a step for culturing the microorganism.

The invention provides a preparation process of high-purity D-panthenol, and belongs to the technical field of pharmaceutical chemistry synthesis. The preparation process comprises the following steps: mixing D-pantoic acid lactone and 3-aminopropanol in a solvent for reaction to obtain a D-panthenol crude product, adding an extracting agent into the D-panthenol crude product, carrying out extraction operation for 0-4 times at the temperature of 30-40 DEG C, separating out the D-panthenol crude product, dissolving the D-panthenol crude product with methanol, concentrating until the solvent residue accounts for 15-25% of the mass of the material, and distilling by using a molecular distiller to obtain the D-panthenol. The high-purity D-panthenol is obtained. The process provided by the invention can effectively control the content of 3-aminopropanol, D-pantoic acid lactone and other impurities in D-panthenol, and can ensure the limit level of the impurities in the stable placement period of the product. The method disclosed by the invention is strong in impurity removal capacity, high in distillation efficiency, simple and convenient to operate, good in product quality, high in stability and suitable for large-scale production.

The present invention relates to the use of PanB from Modestobacter versicolor in the production of pantothenate or pantoic acid and in the preparation of microorganisms producing pantothenate or pantoic acid and methods of producing pantothenate or pantoic acid using the same.

The invention discloses an I-type CRISPRi (clustered regularly interspaced short palindromic repeats i) system based on ComQXPA quorum sensing, an engineering bacterium and an application of the I-type CRISPRi system. According to the invention, a pantoic acid synthesis route is enhanced, cofactors and energy supply are optimized, precursor pyruvic acid accumulation and glucose uptake efficiency are improved, and high-yield D-pantothenic acid engineering chassis bacteria are constructed; and the bacillus subtilis endogenous QS-mediated I-type CRISPRi system is used for dynamic regulation and control of ccpA, odhA and ptsG genes to obtain the probiotic engineering bacteria capable of responding to the population density. The strain can dynamically regulate and balance intracellular metabolic flux through self-induction, intermediate product accumulation is avoided, efficient synthesis of D-pantothenic acid is achieved, the shake flask yield reaches 8.66 g / L, and the yield of a 5L bioreactor reaches 32.26 g / L.

The present disclosure provides: a microorganism introduced with a foreign lipoate metabolic gene; a composition for producing at least one selected from the group consisting of pantothenic acid, pantoic acid, and tryptophan, the composition comprising the microorganism; and a method for producing at least one selected from the group consisting of pantothenic acid, pantoic acid, and tryptophan, the method comprising a step for culturing the microorganism.

The invention provides a high-yield D-pantothenic acid gene engineering bacterium based on an ilvY activation system as well as a construction method and application thereof, and belongs to the technical field of gene engineering. According to the invention, high-activity mutants ilvYR257C and S264C are obtained mainly by constructing a fluorescence activated expression system, constructing an ilvY mutant library by using error-prone PCR (Polymerase Chain Reaction) and performing high-throughput screening; the structural stability and binding capacity of the mutant are verified through molecular dynamics simulation; the mutant is integrated into an original strain, and a high-yield genetically engineered bacterium is constructed by combining enhanced expression of a pantoic acid path key gene alsS and inhibition of a D-pantothenic acid degradation path. The genetically engineered bacterium is subjected to fed-batch fermentation in a 5L fermentation tank for 86h, the yield of D-pantothenic acid reaches 137.8 g / L and is increased by 22.5% compared with an original strain, and the yield is 1.64 g / L / h. By optimizing an ilvY activation system, precise regulation and control of a pantoic acid synthesis path are realized, and technical support is provided for industrial efficient production of D-pantothenic acid.

The present application provides an L-pantoic acid lactone dehydrogenase derived from Skermania sp. ID1734, a nucleic acid molecule encoding the L-pantoic acid lactone dehydrogenase, a vector for expressing the L-pantoic acid lactone dehydrogenase in a host cell, and an engineered bacterium expressing the enzyme. The present application further provides an engineered bacterium co-expressing the L-pantoic acid lactone dehydrogenase and any one or more selected from the group consisting of a ketopantoate reductase and a glucose dehydrogenase. The present application further provides a method for producing D-pantoic acid lactone using an engineered bacterium expressing the L-pantoic acid lactone dehydrogenase derived from Skermania sp. ID1734. One of the technical solutions provided by the present application is to provide a recombinant vector or a combination of recombinant vectors, wherein the recombinant vector or the combination of recombinant vectors comprises a gene expressing the L-pantoic acid lactone dehydrogenase. The present application also provides the use or application of the L-pantoic acid lactone dehydrogenase, the nucleic acid molecule, the engineered bacterium, the vector or the combination of vectors in the synthesis of D-pantoic acid lactone.

The invention provides a preparation method of efficient immobilized D-pantoic acid lactone hydrolase. The method comprises the following steps: mixing wet thalli of D-pantoic acid lactone hydrolase with a glutaraldehyde aqueous solution, and pre-crosslinking to obtain a thalli suspension; dissolving sodium alginate in water to obtain a sodium alginate aqueous solution; mixing the thallus suspension with a sodium alginate aqueous solution, and performing emulsification and vacuum degassing treatment to obtain an emulsion; dropwise adding the emulsion into solidification liquid containing calcium chloride and a synergistic cross-linking agent, solidifying, collecting immobilized enzyme microspheres, and washing to obtain the efficient immobilized D-pantoic acid lactone hydrolase. The immobilized D-pantoic acid lactone hydrolase is high in mechanical strength and good in enzyme activity stability, can be repeatedly used for more than 500 times, is applied to catalysis of hydrolysis of DL-pantoic acid lactone, and can realize efficient and low-cost green synthesis of D-pantoic acid.

The present application provides a kind of derived from Rhodococcus erythropolis L-pantoic acid lactone dehydrogenase of (Rhodococcus erythropolis) and its mutant, nucleic acid molecule for encoding the L-pantoic acid lactone dehydrogenase or its mutant, vector for expressing the L-pantoic acid lactone dehydrogenase or its mutant in host cell and the engineering bacteria of expression the enzyme or its mutant.The present application further provides a kind of co-expression the L-pantoic acid lactone dehydrogenase or its mutant and any one or more selected from the following group: molecular chaperone, D-ketopantoate reductase and glucose dehydrogenase.The present application further provides a kind of D-pantoic acid lactone production method, which uses the engineering bacteria of expression the L-pantoic acid lactone dehydrogenase or its mutant derived from Rhodococcus erythropolis .

The present application provides a L-pantoic acid lactone dehydrogenase derived from Rhodococcus erythropolis, and a mutant thereof, a nucleic acid molecule encoding the L-pantoic acid lactone dehydrogenase or the mutant thereof, a vector for expressing the L-pantoic acid lactone dehydrogenase or the mutant thereof in a host cell, and an engineered bacterium expressing the enzyme or the mutant thereof. The present application further provides an engineered bacterium co-expressing the L-pantoic acid lactone dehydrogenase or the mutant thereof and any one or more selected from the group consisting of a chaperone, a D-ketopantoic acid lactone reductase, and a glucose dehydrogenase. The present application further provides a method for producing D-pantoic acid lactone using the engineered bacterium expressing the L-pantoic acid lactone dehydrogenase derived from Rhodococcus erythropolis or the mutant thereof.