39 results about "Acetyl Coenzyme A Synthetase" patented technology

The present invention relates to a method of identifying a heterologous polypeptide having enzymatic activity for converting pyruvate, acetaldehyde or acetate into acetyl-CoA in (the cytosol of) a yeast cell comprising: a) providing a mutated yeast cell comprising a deletion of at least one gene of the (PDH) by-pass, selected from the genes encoding the enzymes pyruvate decarboxylase (PDC), acetaldehyde dehydrogenase (ALD), and acetyl-CoA synthetase (ACS); b) transforming said mutated yeast cell with an expression vector comprising a heterologous nucleotide sequence encoding a candidate polypeptide having potential enzymatic activity for converting pyruvate, acetaldehyde or acetate into acetyl-CoA; c) testing said recombinant mutated yeast cell for its ability to grow on minimal medium containing glucose as sole carbon source, and d) identifying said candidate polypeptide as a heterologous polypeptide having enzymatic activity for converting pyruvate, acetaldehyde or acetate into acetyl-CoA in (the cytosol of) said yeast cell when growth of said cell is observed. The invention further relates to a method of producing a fermentation production such as butanol.

The invention discloses a method for producing eriodictyol by reforming escherichia coli in metabolic engineering, belonging to the field of metabolic engineering. The step of transforming naringenin into the eriodictyol is successfully realized through a gene engineering technology by carrying out fusion expression on two genes, namely a flavone 3' hydroxylase gene tF3'H with a membrane binding sequence cut and a P450 reductase gene tCPR, in escherichia coli. A tyrosine ammonia lyase gene TAL from R.glutinis, 4-coumarate:coenzyme A ligase gene 4CL from P.crispum, a chalcone synthase gene CHS from P.hybrida and a chalcone isomerase gene CHI from M.sativa are simultaneously expressed to realize that naringenin is directly produced through tyrosine, so that an engineered strain for directly producing eriodictyol through tyrosine is established. According to the method, in order to increase the output of eriodictyol, an acetokinase ackA gene contained in an escherichia coli genome is removed by being knocked, an acetyl coenzyme A synthetase gene acs and an acetyl coenzyme A carboxylase gene ACC are excessively expressed, and the output of eriodictyol can finally reach 106.7 mg/L.

The invention discloses engineering bacteria for producing trans-4-hydroxy-L-proline and a construction method and an application thereof. The construction method of the engineering bacteria provided by the invention includes the steps of A1) and A2): A1) introducing a L-proline-4-hydroxylase gene, a glutamate-5-kinase gene and a glutamate-5-semialdehyde dehydrogenase gene into a receptor cell; and A2) knocking an alpha-ketoglutaricdehydrogenase gene, an isocitratlyase gene or a proline dehydrogenase gene of the receptor cell out, or replacing a pyruvic oxidase gene of the receptor cell with an acetylcoenzyme A synthetase gene; and carrying out a reaction of the recombinant cell and a substrate to obtain the trans-4-hydroxy-L-proline. Experiments prove that the production method of the trans-4-hydroxy-L-proline can be used for production of the trans-4-hydroxy-L-proline.

The invention discloses a gene engineering strain capable of synthesizing phloroglucinol from acetic acid and a construction method and application thereof and belongs to the technical field of genetic engineering. The gene engineering strain disclosed by the invention co-expresses an acetyl coenzyme A synthetase gene acs, a malate dehydrogenase gene maeB, a polyketene anhydride synthetase gene phlD and a multi-resistance factor gene marA. Meanwhile, the invention further provides a construction method of the gene engineering strain. The gene engineering strain disclosed by the invention can be used for synthesizing phloroglucinol from acetic acid and has the characteristics of low production cost and high yield, the yield can reach over 1.2g/L, and the maximum output can reach 30% and is 52% of the theoretical yield, so that the gene engineering strain is suitable for industrialized popularization.

The invention relates to a free fatty acid determination reagent kit which comprises dual reagents. The reagent 1 comprises Good's buffer solution (pH(potential of hydrogen)=7.0), coenzyme A, ATP (adenosine triphosphate), acetylcoenzyme A synthetase, magnesium chloride (MgCL2), Trinder bound constituent, tween 80 and liquid biological preservative/Proclin 300, and the reagent 2 comprises Good's buffer solution (pH=7.0), acetylcoenzyme A oxidase (ACOD), peroxidase (POD), Trinder bound constituent, tween 80 and liquid biological preservative/Proclin 300. The free fatty acid determination reagent kit has the remarkable advantages that the reagent kit is directly used for an automatic biochemical analyzer and is simple; and the adopted automatically separated and purified acetylcoenzyme A synthetase and acetylcoenzyme A oxidase (ACOD) have high specificity on free fatty acid in serum.

Cell that is genetically modified comprising:

• a) one or more nucleotide sequence encoding a NAD+-dependent acetylating acetaldehyde dehydrogenase (E.C. 1.2.1.10);
• b) one or more nucleotide sequence encoding a acetyl-CoA synthetase (E.C. 6.2.1.1);
• c) one or more nucleotide sequence encoding a glycerol dehydrogenase (E.C. 1.1.1.6); and
• d) one or more nucleotide sequence encoding a dihydroxyacetone kinase (E.C. 2.7.1.28 or E.C. 2.7.1.29).

The invention discloses a genetic engineering bacterium for producing glycollic acid by acetic acid, and a building method and application thereof. A method for preparing the genetic engineering bacterium for producing glycollic acid provided by the invention comprises the following steps of improving the expression and/or activity of acetylcoenzyme A synthetase, phosphotransacetylase, acetokinase, citrate synthase, isocitrate lyase, isocitrate dehydrogenase kinase and glyoxylate reductase in the recipient bacterium; reducing the expression and/or the activity of malate synthase, glycolate oxidase and isocitrate lyase repressor protein in the recipient bacterium, so that the engineering bacterium for producing the glycollic acid is obtained. The recipient bacterium is a bacterium capable of using acetic acid as a carbon source for growth. The prepared genetic engineering bacterium uses acetic acid as the carbon source for producing the glycollic acid; in addition, the production quantity and yield of the glycollic acid in shaking culture can reach the relatively high level; relatively good industrial application prospects are realized.

The invention belongs to the technical field of biological medicine, and specifically provides a human pathogen clostridium difficile inhibitor taking acetylcoenzyme A synthetase (ACS) as a target. This invention utilizes a fast reaction kinetics technique to successfully find an acetylcoenzyme A synthetase inhibitor with high inhibitory activity, which comprises 1,10-o-phenanthroline, 2,2-bipyridyl and 8-hydroxyquinoline, wherein the conventional clinically applied medicament, namely 8-hydroxyquinoline, can inhibit methyl transfer activity of acetylcoenzyme A nearly by 100 percent in a low concentration. A method of the invention is the only reported method at present for an efficient inhibitor of human pathogen clostridium difficile acetylcoenzyme A synthetase. The inhibitor is of important application value in the field of biological medicine.

The invention designs a naringenin in-vitro enzymatic synthesis method based on malonyl coenzyme A regeneration. On the basis of constructing recombinant expression plasmids of an acetyl coenzyme A synthetase ACS gene and an acetyl coenzyme A carboxylase ACC1 gene, the recombinant plasmids are respectively transformed into escherichia coli and yeast cells to express a target protein, and the purified ACS and ACC1 recombinant proteins are added into a naringenin in-vitro enzymatic synthesis system to realize the regeneration of malonyl coenzyme A and the in-vitro low-cost enzymatic synthesis of naringenin by using 4-coumaric acid as a substrate. The invention designs a new reaction system, which can be regenerated and utilized without adding expensive malonyl coenzyme A and finally generates naringenin.

The present invention relates to Saccharomyces sp. capable of producing lactic acid with a decreased activity of pyruvate decarboxylase (PDC) and increased activities of aldehyde dehydrogenase (ALD) and acetyl-CoA synthetase (ACS), and a method of producing lactic acid from the culture medium obtained by culturing the microorganism.

The invention discloses recombination acetyl coenzyme A synthetase. The amino acid sequence of therecombination acetyl coenzyme A synthetase is shown in SEQ ID No.2 and the nucleotide sequence of the recombination acetyl coenzyme A synthetase is shown in SEQ ID NO.1.The recombination acetyl coenzyme A synthetase has the characteristics of high thermal stability and high activity; the retaining rate of enzyme activity is 59.6% after the recombination acetyl coenzyme A synthetase is treated for 30 min at the temperature of 60 DEG C and is much improved compared with constitutive enzyme; an apparent Km value of a catalyzed oleic acid acetylation reaction is 1.2*10<-5>M; the thermal stability of enzyme in a kit can be improved, and temperature influences can be avoided or reduced in the transportation and utilization processes of the kit.

The invention discloses a method for enhancing utilization of acetic acid and improving L-arginine produced by escherichia coli fermentation, and belongs to the technical field of bioengineering. Acetobacter pasteurianus sourced acetyl coenzyme A synthetase ACS is successfully expressed on a pET28a carrier, and then a recombinant plasmid is transferred into modified escherichia coli ARG-4. The recombinant escherichia coli is inoculated into a fermentation culture medium containing glucose and is subjected to shake flask fermentation for 48 hours, so that the yield of the L-arginine in the fermentation liquid can reach 7.51 g/L, and meanwhile, the content of a byproduct acetic acid is 2.53 g/L. The recombinant Escherichia coli is inoculated into a fermentation medium containing glucose and is fermented in a fermentation tank for 48 hours, so that the yield of L-arginine in fermentation liquor can reach 15.07 g/L, meanwhile, the content of byproduct acetic acid is only 0.41 g/L. In the whole fermentation process, acetic acid can be effectively utilized to generate acetyl coenzyme A, and the recombinant Escherichia coli has the advantages of low raw material cost, few byproducts and easiness in operation.

The invention discloses an engineering bacterium for biosynthesizing resveratrol by taking L-tyrosine as a substrate, construction and application, and belongs to the technical field of biology. According to the invention, a simple and effective implementation method is provided for safe biotransformation of the resveratrol; biosynthesis from the L-tyrosine to the resveratrol is implemented in cells by adopting co-expression on TAL, 4CL and STS modules; the content of the product resveratrol is remarkably increased by expressing an acetyl coenzyme A carboxylase gene and an acetyl coenzyme A synthetase gene; and the yield of the resveratrol biotransformed by the engineering bacterium is as high as 5.5-6 g/L.

The invention discloses a method for improving bacitracin produced by bacillus licheniformis and belongs to the technical field of microorganisms. The bacillus licheniformis ATH with high yield of bacitracin is screened from a feed sample, the acetyl coenzyme A synthetase Bsacs gene derived from bacillus subtilis is expressed in the bacillus licheniformis ATH through a genetic engineering method, supply of amino acid formed by intracellular bacitracin is increased, and the titer of bacitracin is improved. After the obtained recombinant bacillus licheniformis is inoculated into a fermentation culture medium containing soybean meal for fermentation culture for 42 hours, the titer of the obtained bacitracin is 1928 IU/mL and is improved by 46.16% compared with that of the bacitracin of an original strain ATH, and meanwhile, the content of bacitracin A is 1081 IU/mL and is improved by 37.2% compared with that of the original strain. The bacitracin production efficiency in the whole fermentation process is high, the raw material cost is low, and the method is simple and easy to operate.

The invention relates to an inorganic phosphorus (phosphate radical) diagnosis/determination kit using the technology of an enzyme colorimetric method and an ELISA method, a method for determining inorganic phosphorus (phosphate radical) concentration and composition and components of a reagent, and belongs to the technical field of test and determination of medical science, food, and environment. The kit comprises the following main components: buffer solution, coenzyme, adenoside, bisphosphate, acetyl coenzyme A, pyruvic acid, 5'nucleotidase, acetyl-coA-synthetase, pyruvate dehydrogenase and a stabilizer. The method comprises the following steps: mixing a sample and the reagent according to a certain volume ratio to perform a series of enzymic reaction, and then placing reactants under a UV/ visible light analyzer to test the ascending speed of absorbance at the position where the dominant wave length is 340nm to determine the inorganic phosphorus (phosphate radical) concentration.

The invention relates to a human glucagon-like peptide-1 receptor (GLP-1R) activator and application thereof. The human glucagon-like peptide-1 receptor activator comprises a compound shown in a formula I which is described in the specification or a pharmaceutically acceptable salt of a compound, and R1, R2, R3, R4, R5 and R6 are respectively and independently selected from any one of H, C1-C4 alkyl, sulfydryl, phosphate group, vinyl or acetyl. It is found for the first time that the compound shown in the formula I or the pharmaceutically acceptable salt thereof can improve glycolipid metabolism of whole-body tissue cells, enhance oxidative degradation of lipid molecules, inhibit expression of adipogenic genes, inhibit activity of acetyl coenzyme A synthetase and reduce synthesis of triglyceride, glucose-induced insulin synthesis and secretion are improved, insulin antagonism is relieved, compensatory amplification of pancreatic beta cells is relieved, and the survival rate and functions of pancreatic cells in the II-type diabetes mellitus-like state are improved.

The invention relates to an inorganic phosphorus (phosphate radical) diagnosis/determination kit using the technology of an enzyme colorimetric method and an ELISA method, a method for determining inorganic phosphorus (phosphate radical) concentration and composition and components of a reagent, and belongs to the technical field of test and determination of medical science, food, and environment. The kit comprises the following main components: buffer solution, reduced coenzyme, adenoside, bisphosphate, acetyl coenzyme A, 5'nucleotidase, acetyl-coA-synthetase, aldehyde dehydrogenase, alcohol dehydrogenase and a stabilizer. The method comprises the following steps: mixing a sample and the reagent according to a certain volume ratio to perform a series of enzymic reaction, and then placing reactants under a UV/ visible light analyzer to test the descending speed of absorbance at the position where the dominant wave length is 340nm to determine the inorganic phosphorus (phosphate radical) concentration.