61 results about "Malonyl Coenzyme A" patented technology

Disclosed are novel acyl-CoA synthetases and novel acyltransferases, nucleic acid molecules encoding the same, recombinant nucleic acid molecules and recombinant host cells comprising such nucleic acid molecules, genetically modified organisms (microorganisms and plants) comprising the same, and methods of making and using the same. Also disclosed are genetically modified organisms (e.g., plants, microorganisms) that have been genetically modified to express a PKS-like system for the production of PUFAs (a PUFA PKS system or PUFA synthase), wherein the organisms have been modified to express an acyl-CoA synthetase, to express an acyl transferase, to delete or inactivate a fatty acid synthase (FAS) expressed by the organism, to reduce competition for malonyl CoA with the PUFA synthase or to increase the level of malonyl CoA in the plant or plant cell, and in one aspect, to inhibit KASII or KASIII. Additional modifications, and methods to make and use such organisms, in addition to PUFAs and oils obtained from such organisms, are disclosed, alone with various products including such PUFAs and oils.

This invention relates to metabolically engineered microorganism strains, such as bacterial strains, in which there is an increased utilization of malonyl-CoA for production of a chemical product, which includes 3-hydroxypropionic acid.

The present invention provides methods for the use of compounds as depicted by structure I, pharmaceutical compositions containing the same, and methods for the prophylaxis, management and treatment of metabolic diseases and diseases modulated by MCD inhibition. The compounds disclosed in this invention are useful for the prophylaxis, management and treatment of diseases involving in malonyl-CoA regulated glucose / fatty acid metabolism pathway. In particular, these compounds and pharmaceutical composition containing the same are indicated in the prophylaxis, management and treatment of cardiovascular diseases, diabetes, cancer and obesity.

Provided is a screening method for compounds affecting fatty acid biosynthesis, the method comprising: providing a reaction mixture comprising: an acyl carrier moiety or enzymes and precursors sufficient to generate the acyl carrier moiety; a bacterial enzymatic pathway comprising at least two consecutively acting enzymes selected from the group consisting of: malonyl-CoA:ACP transacylase, beta-ketoacyl-ACP synthase III, NADPH dependent beta-ketoacyl-ACP reductase, beta-hydroxylacyl-ACP dehydrase and enoyl-ACP reductase; and substrates and cofactors required for the operation of the enzymes; contacting the reaction mixture with a prospective bioactive agent; conducting a high throughput measurement of the activity of the enzymatic pathway; and determining if the contacting altered the activity of the enzymatic pathway.

Disclosed are novel acyl-CoA synthetases and novel acyltransferases, nucleic acid molecules encoding the same, recombinant nucleic acid molecules and recombinant host cells comprising such nucleic acid molecules, genetically modified organisms (microorganisms and plants) comprising the same, and methods of making and using the same. Also disclosed are genetically modified organisms (e.g., plants, microorganisms) that have been genetically modified to express a PKS-like system for the production of PUFAs (a PUFA PKS system or PUFA synthase), wherein the organisms have been modified to express an acyl-CoA synthetase, to express an acyl transferase, to delete or inactivate a fatty acid synthase (FAS) expressed by the organism, to reduce competition for malonyl CoA with the PUFA synthase or toincrease the level of malonyl CoA in the organism, and in one aspect, to inhibit KASII or KASIII. Additional modifications, and methods to make and use such organisms, in addition to PUFAs and oils o btained from such organisms, are disclosed, alone with various products including such PUFAs and oils.

This invention relates to a recombinant microorganism capable of producing isoprene and isoprene production with the use of such recombinant microorganism with good efficiency. In this invention, the acetoacetyl-CoA synthase gene encoding an enzyme capable of synthesizing acetoacetyl-CoA from malonyl-CoA and acetyl-CoA and one or more genes involved in isoprene biosynthesis that enables synthesis of isoprene from acetoacetyl-CoA are introduced into a host microorganism.

A microbial cell is used for producing at least one fatty acid ester, wherein the cell is genetically modified to contain (i) at least one first genetic mutation that enables the cell to produce at least one fatty acid and / or acyl coenzyme A (CoA) thereof by increased enzymatic activity in the cell relative to the wild type cell of malonyl-CoA dependent and malonyl-ACP independent fatty acyl-CoA metabolic pathway, wherein the fatty acid contains at least 5 carbon atoms; and (ii) a second genetic mutation that increases the activity of at least one wax ester synthase in the cell relative to the wild type cell and the wax ester synthase has sequence identity of at least 50% to a polypeptide of SEQ ID NO: 1-8 and combinations thereof or to a functional fragment of any of the polypeptides for catalyzing the conversion of fatty acid and / or acyl coenzyme A thereof to the fatty acid ester.

The long-term usage of AICR (5-aminio, 4-imidazole carboxamide riboside) to produce sustained metabolic and biological changes in mammals that overcome insulin resistance, i.e., increase insulin sensitivity, and result in benefits in diseases and conditions such as diabetes, hypertension, atherosclerosis, polycystic ovary syndrome and gallstones is described long-term usage of AICAR, particularly intermittent administration, e.g., three days per week, appears to have some of the positive effects of exercise, having an impact on the amount Of food consumed by a subject and resulting in reduced fat build-up and increase in muscle mass. Therefore, AICAR administration has a positive impact in reducing obesity. AICAR can also Prove useful in preventing or treating vascular diseases associated with hyperglycemia, high plasma levels of free fatty acids (FFA) and triglyceride, and insulin resistance by virtue of the fact that this agent activates fatty acid oxidation. Animal tests have Shown that chronic intermittent treatment with AICAR has not resulted in any noticeable toxic effects. AICAR and related compounds are activators of AMP-activated protein kinase (AMPK) and, furthermore, are effective at decreasing malonyl CoA levels in the animal.

The invention discloses an extract of yangtao actinidia root and a preparation method and applications thereof. The extract of yangtao actinidia root, which is provided by the invention, is prepared by the following method: using water and / or an organic solvent as an extractant to extract the extract of yangtao actinidia root from yangtao actinidia root. Experiment shows that the extract of yangtao actinidia root can effectively inhibit the activity of fatty acid synthase. The inhibition kinetic analysis shows that the inhibition of the extract of yangtao actinidia root for the fatty acid synthase and the substrates of acetylcoenzyme A and NADPH have a competitive and non-competitive mixed type relationship, but more close to the non-competitive relationship; and the inhibition of the extract of yangtao actinidia root for the fatty acid synthase and the substrate of malonyl coenzyme A have a competitive and non-competitive mixed type relationship. The extract of yangtao actinidia root can be applied to the preparation of weight-reducing medicament and the medicament for preventing and treating cancers, and also can be widely applied as food, health product and daily chemical additive.

The invention discloses related biological materials of fusion protein Nt4CL3aPcSTS and application of the related biological materials. The related biological materials of the fusion protein Nt4CL3aPcSTS provided by the invention comprise the fusion protein Nt4CL3aPcSTS, wherein the fusion protein Nt4CL3aPcSTS is obtained by connecting protein Nt4CL and protein PcSTS through three amino acid residues, and the protein Nt4CL is a protein constituted by 1st-542nd amino acid residues from the Nth end of amino acid sequence SEQ ID No.1; the protein PcSTS is a protein showed by the 546th-933rd amino acid residues from the Nth end of amino acid sequence SEQ ID No.1. The related biological materials of the fusion protein Nt4CL7aPcSTS can be used for catalyzing 4-cumaric acid and malonyl coenzyme A to generate resveratrol with bioactivity.

The invention provides an engineering strain with high yield of FK520 (ascomycin). A gene expression cassette is integrated in a genome of the strain, and the gene expression cassette expresses a protein encoded by a biosynthetic gene of ethyl malonyl coA. The yield of FK520 can be increased substantially by the transformed strain, and the proportion (mass ratio) of the main product FK520 to sideproduct FK523 is increased.

The invention relates to a 1-propyl-alcohol production microorganism introducing a heterologous metabolic pathway and a method for producing 1-propyl-alcohol by means of the microorganism. A metabolic pathway of 1-propyl-alcohol production is introduced into the 1-propyl-alcohol microorganism and generation of by-product lactate is prevented. Specifically, the gene adhE2 of Alcohol dehydrogenase is efficiently expressed in escherichia coli, the gene epi of Methylmalonyl-CoA epimerase is encoded, the gine Sbm of Methylmalonyl CoA mutase is encoded, and the gene ygfG of Methylmalonyl-CoA decarboxylase and the gene mhpF of Acetaldehyde dehydrogenase 2 are encoded. The genes are built on expression plasmids provided with T1 promoters and antibiotic resistance. The built plasmids are transferred to an escherichia coli microorganism, a production strain capable of producing 1-propyl-alcohol by means of glucose is obtained, meanwhile, a large amount of the by-product lactate is obtained, and the method for producing 1-propyl-alcohol is more effective by preventing generation of the by-product.

The invention discloses a synechocystis-6803 genetically engineered bacterium capable of producing 3-hydroxypropionic acid, and a construction method and application thereof. The construction method is as follows: a malonyl coenzyme A reductase gene in orange chloroflexus is cloned into synechocystis 6803; and acetylcoenzyme A carboxylase, biotin acylase and an NAD(P) transhydrogenase gene in the synechocystis 6803 are expressed. The synechocystis 6803 is transformed through a synthetic biology method to obtain the synechocystis-6803 genetically engineered bacterium capable of producing the 3-hydroxypropionic acid. The experiments prove that the final yield of the 3-HP (3-hydroxypropionic acid) of the genetically engineered bacterium is up to 837.18mg / L, which is of great theoretical and practical significances for the production of the 3-HP through photosynthetic microorganisms.

This invention relates to metabolically engineered microorganism strains, such as bacterial strains, in which there is an increased utilization of malonyl-CoA for production of a chemical product, which includes polyketides and 3-hydroxypropionic acid.

The invention provides a streptomycete strain mutagenesis preparation method for high-yield production of tacrolimus. According to the invention, mutagenesis breeding is performed with nitrosoguanidine (NTG), and then active substances of vibralactone and ethyl malonyl coenzyme A are used as selection pressures to achieve the aim of increasing the yield of tacrolimus. According to the invention, the yield of tacrolimus prepared by taking the active substances of vibralactone and ethyl malonyl coenzyme A synergistically as selection pressures is obviously increased compared with the yield of mutagenesis breeding by using a single selection pressure. Streptomyces tsukubaensis prepared by taking the active substances of vibralactone and ethyl malonyl coenzyme A as selection pressures has thecharacteristics of high activity of ethyl malonyl coenzyme A and high activity of lipase, and the yield of tacrolimus is improved due to high activity of ethyl malonyl coenzyme A and high activity oflipase, and generation of by-products is reduced. The yield of combined use of vibralactone and ethyl malonyl coenzyme A is obviously higher than the yield of mutagenesis breeding by using a single selection pressure.

A system for carbon fixation is provided. The system comprises enzymes which catalyze reactions of a carbon fixation pathway, wherein at least one of the reactions of the carbon fixation pathway is a carboxylation reaction, wherein products of the reactions of the carbon fixation pathway comprise oxaloacetate and malonyl-CoA, wherein an enzyme which performs the carboxylation reaction is selected from the group consisting of phophoenolpyruvate (PEP) carboxlase, pyruvate carboxylase and acetyl-CoA carboxylase and wherein an export product of the carbon fixation pathway is glyoxylate. Additional carbon fixation pathways are also provided and methods of generating same.

The invention discloses a molecular probe for detecting malonyl coenzyme A. The moklecular probe contains fusion protein of NanoLuc luciferase large subunit, FapRdelta43 protein and NanoLuc luciferasesubunit from the N end to the C end. The molecular probe can both detect the malonyl coenzyme A in the extract of sample in vitro and detect and perform detection and dynamic observation on the malonyl coenzyme A in living cells; most importantly, the malonyl coenzyme A can be detected in different organelles of the living cells, simpleness in operation is achieved, and therefore, the fusion protein has the potential to become a commercial product.

A yeast cell having a reduced level of activity of NAD dependent glyceraldehyde-3-phosphate dehydrogenase (GAPDH) has at least one exogenous gene encoding NADP dependent GAPDH and / or has up-regulation of at least one endogenous gene expressing NADP dependent GAPDH, wherein combined expression of the enzymes NADP dependent GAPDH, PDC, ALD, ACS, ACC* and MCR in said host cell increases metabolic flux towards 3-HP via malonyl-CoA compared to an otherwise similar yeast cell lacking said genetic modification.

This invention relates to metabolically engineered microorganism strains, such as bacterial strains, in which there is an increased utilization of malonyl-CoA for production of a fatty acid or fatty acid derived product, wherein the modified microorganism produces fatty acyl-CoA intermediates via a malonyl-CoA dependent but malonyl-ACP independent mechanism.

The invention provides a technical method using 'photosynthetic bacteria', namely blue algae as the substrate organism to synthesize phloretin. The method mainly includes: exogenously expressing 4 coumarate Coenzyme A Ligase (4CL) and Chalcone synthase (CHS) which are two key enzymes for phloretin synthesis in the blue algae so as to use p-hydroxybenzene propanoic acid (also called phloretic acid)as the substrate to catalytically synthesize the phloretin, the 4CL catalyzes single-molecule p-hydroxybenzene propanoic acid to generate p-hydroxybenzene propionyl coenzyme A, the CHS catalyzes three-molecule malonyl coA and the single-molecule p-hydroxybenzene propionyl coenzyme A to synthesize the single-molecule phloretin. The method has the advantages that the phloretin is produced by usingone microorganism which can perform photosynthesis, the phloretin biological synthesizing approach which is cheap in raw material, simple in equipment, low in environment pollution and high in yield is created, and the method conforms to the direction of green production.

Organic acid-tolerant microorganisms and methods of using same. The organic acid-tolerant microorganisms comprise modifications that reduce or ablate AcsA activity or AcsA homolog activity. The modifications increase tolerance of the microorganisms to such organic acids as 3-hydroxypropionic acid (3HP), acrylic acid, and propionic acid. Further modifications to the microorganisms such as increasing expression of malonyl-CoA reductase and / or acetyl-CoA carboxylase provide or increase the ability of the microorganisms to produce 3HP. Methods of generating an organic acid with the modified microorganisms are provided. Methods of using acsA or homologs thereof as counter-selectable markers include replacing acsA or homologs thereof in cells with genes of interest and selecting for the cells comprising the genes of interest with amounts of organic acids effective to inhibit growth of cells harboring acsA or the homologs.

The invention discloses a gene engineering bacteria with high-yield malonyl coenzyme A and a construction method and application thereof. The gene engineering bacteria is constructed by: knocking outfive genes (ldhA, pta, frdA, poxB and adhE) in the Escherichia coli genome, and then introducing malonyl coenzyme A synthesis pathway genes including acetyl coenzyme A synthetase gene of the Escherichia coli, acetyl coenzyme A carboxylase gene of Salmonella enteritidis and biotin ligase gene of Corynebacterium glutamicum. According to the gene engineering bacteria of the invention, a highly-efficient accumulation of the malonyl coenzyme A can be realized by inhibiting internal acetyl coenzyme A outflow pathways of the engineering bacteria and constructing malonyl coenzyme A synthesis pathwaysinto different expression vectors to be transferred into the engineering bacteria; the engineering bacteria can efficiently synthesize a precursor, malonyl coenzyme A, of flavonoid compounds by takingacetic acid, a metabolite by-product of the Escherichia coli, as a substrate; and the engineering bacteria can be used to increase the yield of naringenin, a skeleton precursor of the flavonoid compounds, synthesized by a microbiological method.

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 invention discloses a method for promoting recombinant yarrowia lipolytica bacteria to synthesize phloretin. According to the method, tyrosine is used as a raw material, recombinant yarrowia lipolytica bacteria are used as host bacteria, and the phloretin is obtained through the catalytic reaction of a plurality of enzymes in the host bacteria. According to the invention, the high accumulation amount of acetyl coenzyme A in arrowia lipolytica bacterium cells is utilized, and by introducing a beneficial mutant fragment of a POX2 promoter, driving the acetyl coenzyme A to be converted into a key gene ACC1 of malonyl coenzyme A and carrying out high-efficiency overexpression on a synthetic key gene 4CL of p-hydroxybenzene propionyl coenzyme A, the accumulation amounts of two important precursor substances, i.e., the malonyl coenzyme A and the p-hydroxybenzene propionyl coenzyme A, for synthesizing the phloretin are increased, so that the yield of the phloretin in the host bacteria is finally increased. According to the method, a phloretin anabolic pathway is constructed in microorganisms, large-scale extraction and separation processes are effectively avoided, and the method is environment-friendly and pollution-free and meets the new requirements of current green production.

The invention discloses use of a branched chain alpha-keto acid dehydrogenase complex in preparing a malonyl coenzyme A. The invention discloses a method for preparing a malonyl coenzyme A by using abranched chain alpha-keto acid dehydrogenase complex. The method comprises the following steps: introducing a coding gene of the branched chain alpha-keto acid dehydrogenase complex into a biologicalcell and expressing the coding gene of the branched chain alpha-keto acid dehydrogenase complex to obtain a recombinant cell; and culturing the recombinant cell to obtain the malonyl coenzyme A. The branched chain alpha-keto acid dehydrogenase complex is a complete set of proteins composed of M1) or M2): M1): bkdF, bkdG, bkdH and lpdA1; and M2: bkdA, bkdB, bkdC and lpdA1. Experiments prove that the branched chain alpha-keto acid dehydrogenase complex can be used for preparing the malonyl coenzyme A and also can be used for preparing a target product taking the malonyl coenzyme A as an intermediate product.