82 results about "Ketoacid decarboxylase" patented technology

A non-naturally occurring microbial organism having a 3-hydroxypropanoic acid (3-HP) pathway includes at least one exogenous nucleic acid encoding 3-HP pathway enzyme expressed in a sufficient amount to produce 3-HP. The 3-HP pathway includes a 2-keto acid decarboxylase, a CoA-dependent oxaloacetate dehydrogenase, or a malate decarboxylase. A method for producing 3-HP includes culturing a non-naturally occurring microbial organism having a 3-HP pathway that includes at least one exogenous nucleic acid encoding a 3-HP pathway enzyme expressed in a sufficient amount to produce 3-HP under conditions and for a sufficient period of time to produce 3-HP. The 3-HP pathway includes a 2-keto acid decarboxylase, a CoA-dependent oxaloacetate dehydrogenase, or a malate decarboxylase.

The invention relates to the genetic modification of Cyanobacteria for the production of ethanol, and more particularly, to the genetic modification of Cyanobacteria by incorporating the genetic information encoding for pyruvate decarboxylase (pdc) and alcohol dehydrogenase (adh).

A yeast strain, wherein the yeast strain is transformed with at least one copy of a gene coding for <SMALLCAPS>D< / SMALLCAPS>-lactate dehydrogenase functionally linked to a promoter sequence allowing the expression of the gene in the yeast strain and the yeast strain has undergone disruption of one or more pyruvate decarboxylase genes or pyruvate dehydrogenase genes. Also, a method of producing <SMALLCAPS>D< / SMALLCAPS>-lactic acid including culturing such a yeast strain in a medium and recovering <SMALLCAPS>D< / SMALLCAPS>-lactic acid.

Mutant thermophilic organisms that consume a variety of biomass derived substrates are disclosed herein. Strains of Thermoanaerobacterium saccharolyticum with acetate kinase and phosphotransacetylase expression eliminated are disclosed herein. Further, strain ALK1 has been engineered by site directed homologous recombination to knockout both acetic acid and lactic acid production. Continuous culture involving a substrate concentration challenge lead to evolution of ALK1, and formation of a more robust strain designated ALK2. The organisms may be utilized for example in thermophilic SSF and SSCF reactions performed at temperatures that are optimal for cellulase activity to produce near theoretical ethanol yields without expressing pyruvate decarboxylase.

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 subject invention concerns the transformation of Gram-positive bacteria with heterologous genes which confer upon these microbes the ability to produce ethanol as a fermentation product. Specifically exemplified is the transformation of bacteria with genes, obtainable from Zymomonas mobilis, which encode pyruvate decarboxylase and alcohol dehydrogenase.

The present invention concerns a new method for the biological preparation of a diol comprising culturing a microorganism genetically modified for the bioproduction of an aliphatic diol, wherein the microorganism comprises a metabolic pathway for the decarboxylation of a hydroxy-2-keto-aliphatic acid metabolite with an enzyme having a 2-keto acid decarboxylase activity, the product obtained from said decarboxylation step being further reduced into the corresponding aliphatic diol, and wherein the microorganism is genetically modified for the improved production of said hydroxy-2-keto-aliphatic acid metabolite.The invention also concerns a modified microorganism for the production of an aliphatic diol.

The invention provides isolated nucleic acids molecules which encode pyruvate decarboxylase enzymes having improved decarboxylase activity, substrate affinity, thermostability, and activity at different pH. The nucleic acids of the invention also have a codon usage which allows for high expression in a variety of host cells. Accordingly, the invention provides recombinant expression vectors containing such nucleic acid molecules, recombinant host cells comprising the expression vectors, host cells further comprising other ethanologenic enzymes, and methods for producing useful substances, e.g., acetaldehyde and ethanol, using such host cells.

The present invention relates to a transformant, containing a lactate dehydrogenase gene which is introduced into Schizosaccharomyces pombe as a host, in which a part of a gene cluster encoding a pyruvate decarboxylase in the Schizosaccharomyces pombe host is deleted or inactivated.

The invention discloses escherichia coli gene engineering bacteria for four enzyme co-expression. The engineering bacteria is characterized by introducing an L-amino acid oxidase gene, an alpha-ketonic acid decarboxylase gene, an alcohol dehydrogenase gene, and an enzyme gene capable of reducing NAD(P) to NAD(P)H. The invention further discloses a construction method and application of recombinantescherichia coli. The engineering bacteria is applied to biological synthesis of phenylethyl alcohol compounds, has the characteristics of simple operation, low cost, high product synthetic efficiency, and high optical purity, and has bright industrial prospects.

The invention discloses recombinant escherichia coli for producing tyrosol and a construction method and application thereof and belongs to the technical field of bioengineering. The escherichia coliheterologously expresses a saccharomyces cerevisiae pyruvate decarboxylase gene ARO10* after codon optimization. According to the recombinant escherichia coli, the ARO10* gene is integrated while a lacI locus, a trpE locus, a pabB locus, a pabA locus and a pykF locus of an escherichia coli genome are deleted, and a strain containing multiple copies of the ARO10* gene is obtained. On the basis of the recombinant strain, the ARO10* gene is randomly integrated at the multiple loci, and the ARO10* gene is inserted at a yccX locus so that the strain with high tyrosol yield can be obtained. No inducer nor antibiotic is required for fermentation by using this strain. After 48 hours of fermentation, the yield of tyrosol can reach 28 mM.

The invention which relates to the field of genetic engineering provides a constitutive expression cassette of Trichoderma reesei. The expression cassette comprises a foreign target gene, a promoter of a pyruvate decarboxylase gene (Ppdc) of Trichoderma reesei, and a terminator of the pyruvate decarboxylase gene (Tpdc) of Trichoderma reesei, wherein the nucleotide sequence of the Ppdc is represented by SEQ ID NO.1, and the nucleotide sequence of the Tpdc is represented by SEQ ID NO.2. The expression cassette which comprises the sequence of the Ppdc of Trichoderma reesei and the sequence of the Tpdc of Trichoderma reesei can express genes from animals, plant or fungi without induction, and can properly modify expression products, protein expression can be carried out with the highly efficient synthesis and secretion capability of Trichoderma reesei, the protein expression level is high, and excessive foreign proteins in the expression products can be avoided. A recombinant strain of the present invention can highly express xylanase without cellulose, and the xylanase has an important application in the papermaking industry.

The invention discloses genetic engineering bacteria and belongs to the technical field of bioengineering. With the introduction of L-alpha-amino acid transaminase, L-glutamic acid gene, alpha-keto acid decarboxylase gene and alcohol dehydrogenase, the genetic engineering bacteria can be applied to biosynthesis of hydroxytyrosol. The invention also discloses a construction method and application of the recombinant Escherichia coli genetic engineering bacteria. And further, substrate transport is promoted and decomposition of the product is reduced by knocking out or enhancing the expression ofrelated genes on the Escherichia coli genome, and the production efficiency of recombinant bacteria is improved. The method for production of hydroxytyrosol by conversion of the recombinant bacteriahas characteristics of simple operation, low cost and high synthesis efficiency of the product, and has a good industrial prospect.

The invention relates to the field of bioengineering and biotechnologies, and discloses a method for producing 2-phenylethanol under biological catalysis. The method comprises the following steps: adding wet cells of E.coli / Pdh, E.coli / Kdc and E.coli / ADH undergoing induced expression into a biological catalysis system taking L-phenylalanine as a substrate for performing a catalytic reaction; centrifuging after finishing the reaction; extracting supernatant to obtain the 2-phenylethanol. In the method, the L-phenylalanine is taken as the substrate, and recombinant escherichia coli transformed with a phenylalanine dehydrogenase gene, a 2-keto acid decarboxylase gene and an alcohol dehydrogenase gene is added into a reaction system for performing a biological deamination, decarboxylation and reduction three-step catalytic reaction in order to generate a product, namely, phenylethanol; a coenzyme, namely, NAD (Nicotinamide Adenine Dinucleotide) is added once and can be recycled, excess ketonic acid and hydrogen sources do not need to be added, no unnecessary side products are generated, a high substrate transforming rate is achieved, and the yield of the phenylethanol is increased remarkably.

The invention discloses a method of generating 1,2,4-butantriol by the in vitro enzyme reaction and application thereof, and belongs to the technical field of bioengineering. The method provided by the invention comprises the following steps: respectively constructing genetically-engineered bacteria of over-expressed D-xylonic acid anhydrase genes, 2-keto acid decarboxylase genes and alcohol dehydrogenase genes; after carrying out fermentation cultivation on the obtained genetically-engineered bacteria, crushing thalli by ultraviolet waves; collecting crude enzyme fluid; and after carrying out mixed adjustment on concentration of D-xylonic acid anhydrase, 2-keto acid decarboxylase and alcohol dehydrogenase, adding a reaction substrate to synthesize 1,2,4-butantriol. The method provided by the invention realizes synthesis of 1,2,4-butantriol in vitro by controlling the enzyme reaction and using D-xylonic acid as the raw material and has the characteristic of good convenience for enlarging production; and the enlarged yield can reach 5.98g / L.

The invention discloses a method for producing D-1,2,4-butantriol through bio-converting cellulosic hydrolyzate. The method comprises the steps of constructing genes for cloning and expressing 2-keto acid decarboxylase, D-xylitol dehydrogenase, D-xylonic acid dehydrase and D-alcohol dehydrogenase, shifting the constructed genes into cells of host bacteria for knocking out xylose isomerase so as to obtain genetic engineering strains, culturing the genetic engineering strains, inoculating the genetic engineering strains to the cellulosic hydrolyzate, and carrying out fermentation so as to produce the D-1,2,4-butantriol. The method disclosed by the invention is easy and feasible, is high in yield and is applicable to industrialization.

The invention provides isolated nucleic acids molecules which encode pyruvate decarboxylase enzymes having improved decarboxylase activity, substrate affinity, thermostability, and activity at different pH. The nucleic acids of the invention also have a codon usage which allows for high expression in a variety of host cells. Accordingly, the invention provides recombinant expression vectors containing such nucleic acid molecules, recombinant host cells comprising the expression vectors, host cells further comprising other ethanologenic enzymes, and methods for producing useful substances, e.g., acetaldehyde and ethanol, using such host cells.

The invention discloses an engineering bacterium for producing hydroxytyrosol with high efficiency and application of the engineering bacterium, and belongs to the technical field of biological engineering. Genes of L-phenylalaninase dehydrogenase, alpha-keto acid decarboxylase and alcohol dehydrogenase are introduced into the genetic engineering bacterium of the present invention, and the engineering bacterium can be applied to biosynthesis of hydroxytyrosol. The invention also discloses a construction method and application of the genetic engineering bacterium of recombinant Escherichia coli. The method for producing hydroxytyrosol by transforming the recombinant bacterium has the characteristics of simple operation, low cost, high efficiency of product synthesis, and has good industrialization prospects.

Mutant thermophilic organisms that consume a variety of biomass derived substrates are disclosed herein. Strains of Thermoanaerobacterium saccharolyticum with acetate kinase and phosphotransacetylase expression eliminated are disclosed herein. Further, strain ALK1 has been engineered by site directed homologous recombination to knockout both acetic acid and lactic acid production. Continuous culture involving a substrate concentration challenge lead to evolution of ALK1, and formation of a more robust strain designated ALK2. Both organisms produce near theoretical ethanol yields without expressing pyruvate decarboxylase

The invention discloses an engineering bacterium and application thereof in tyrosol production and belongs to the technical field of bioengineering. L-alpha-amino acid transaminase, L-glutamate dehydrogenase, alpha-ketone decarboxylase and alcohol dehydrogenase are introduced into the genetic engineering bacterium, and the engineering bacterium can be applied to biosynthesis of the tyrosol. The invention further discloses a construction method and application of recombinant Escherichia coli bacteria. Further, by knocking out or enhancing related genes on Escherichia coli genome, transport of the substrate is promoted, decomposition of the product is reduced, and the production efficiency of the recombinant bacteria is improved. The method for producing tyrosol by utilizing transformation of the recombinant bacteria disclosed by the invention has the characteristics of being simple in operation, low in cost and high in product synthesis efficiency, and has excellent industrial prospects.

The invention relates to a recombinant escherichia coli, a preparation method and a method for synthesizing 3,4-dihydroxybutyric acid, and belongs to the field of biosynthesis. The recombinant escherichia coli is obtained through knocking out xylose isomerase gene xylA, 2-keto acid aldolase gene yjhH, 2-keto acid aldolase gene yagE and alcohol dehydrogenase gene in escherichia coli, overexpressing xylose dehydrogenase gene and / or 2-keto acid decarboxylase gene, and overexpressing aldehyde dehydrogenases gene. By taking xylose as a basic substrate, a compound substrate can be formed by adding glucose and / or glycerinum on the basis of the xylose, and the 3,4-dihydroxybutyric acid is biologically synthesized through the recombinant escherichia coli; by-products are extremely less, and the formation of the by-products can be interdicted through passage metabolism optimization. The recombinant escherichia coli, the preparation method and the method for synthesizing the 3,4-dihydroxybutyric acid have the advantages that the preparation method is simple, the production cycle is short, the cost is low, later continuous optimizing and transformation are realized, and the good industrial development and application prospect is realized.

The invention discloses genetically engineered bacteria and an application thereof in production of BT (D-1,2,4-butanetriol). The genetically engineered bacteria are novel genetically engineered bacteria which are obtained as follows: a 2-keto acid decarboxylase gene MalC, an xylose dehydrogenase gene XylB, an xylonic acid dehydratase gene YjhG and an alcohol dehydrogenase gene YqhD are constructed, cloned and expressed, the genes are transferred into cells of host bacteria BL21(DE3), genetically engineered bacteria BL21-02 are obtained, and new xylonic acid dehydratase is screened on the basis of the genetically engineered bacteria; the genetically engineered bacteria are subjected to fermenting culture for production of BT. The capability of synthesizing BT from D-xylose can be improvedby screening the provided xylonic acid dehydratase gene CcXylD. The optimal xylonic acid dehydratase gene CcXylD and alpha-keto acid decarboxylase gene KdcA from lactococcus lactis are applied to theproduction process of BT, the optimal strain BL21-15 is obtained, and finally, the BT yield can reach 10.66 g / L.

The invention discloses a creatine kinase isoenzyme detection reagent. The reagent consists of a reagent R1 and a reagent R2 according to the volume ratio of 4 to 1, wherein the reagent R1 consists of an imidazole buffer liquid, glucose, nano particles, N-acetylcysteine, sodium ethylene diamine tetracetate, adenosine diphosphate, cozymase I, ribonucleotide, pyruvate decarboxylase, glucose 6-phosphate dehydrogenase, hexokinase, a goat anti-human CK-M polyclonal antibody and trehalose; the reagent R2 consists of an imidazole buffer liquid, phosphocreatine, sodium dodecyl benzene sulfonate and a preservative. The reagent disclosed by the invention is a creatine kinase isoenzyme detection reagent which is stable, accurate, and high in anti-interference, and has very high clinical application value.

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 present invention concerns a new method for the biological preparation of a diol comprising culturing a microorganism genetically modified for the bioproduction of an aliphatic diol, wherein the microorganism comprises a metabolic pathway for the decarboxylation of a hydroxy-2-keto-aliphatic acid metabolite with an enzyme having a 2-keto acid decarboxylase activity, the product obtained from said decarboxylation step being further reduced into the corresponding aliphatic diol, and wherein the microorganism is genetically modified for the improved production of said hydroxy-2-keto-aliphatic acid metabolite. The invention also concerns a modified microorganism for the production of an aliphatic diol.

A metabolically enhanced cyanobacterial cell for the production of ethanol is provided. The metabolically enhanced cyanobacterial cell for the production of ethanol comprises at least one recombinant gene encoding a pyruvate decarboxylase enzyme (Pdc) converting pyruvate to acetaldehyde, and at least one recombinant gene encoding a first Zn2+ dependent alcohol dehydrogenase enzyme (Adh) converting acetaldehyde to ethanol. The invention also provides a method for producing the metabolically enhanced cyanobacterium, a method for producing ethanol with the metabolically enhanced cyanobacterium, and a method for screening of alcohol dehydrogenase enzyme expressing cyanobacterial strains for the presence of NADPH-dependent native alcohol dehydrogenase enzymes.

The invention provides a genetic engineering blue-green alga for producing ethanol through photosynthesis. The genetic engineering blue-green alga contains exogenous pyruvate decarboxylase genes and ethanol dehydrogenase which are integrated in chromosomes. The invention further provides a construction body, a carrier and a preparation method for preparing the genetic engineering blue-green alga, and a method for producing ethanol by using the genetic engineering blue-green alga.

The invention which relates to the field of genetic engineering provides a constitutive expression cassette of Trichoderma reesei. The expression cassette comprises a foreign target gene, a promoter of a pyruvate decarboxylase gene (Ppdc) of Trichoderma reesei, and a terminator of the pyruvate decarboxylase gene (Tpdc) of Trichoderma reesei, wherein the nucleotide sequence of the Ppdc is represented by SEQ ID NO.1, and the nucleotide sequence of the Tpdc is represented by SEQ ID NO.2. The expression cassette which comprises the sequence of the Ppdc of Trichoderma reesei and the sequence of the Tpdc of Trichoderma reesei can express genes from animals, plant or fungi without induction, and can properly modify expression products, protein expression can be carried out with the highly efficient synthesis and secretion capability of Trichoderma reesei, the protein expression level is high, and excessive foreign proteins in the expression products can be avoided. A recombinant strain of the present invention can highly express xylanase without cellulose, and the xylanase has an important application in the papermaking industry.

The present invention concerns a microorganism genetically modified for the bioproduction of 1,3-propanediol from sucrose, wherein the microorganism comprises: a two-step metabolic pathway for the production of 1,3-propanediol, comprising a first step of decarboxylation of 4-hydroxy-2-ketobutyrate with an enzyme having a 2-keto acid decarboxylase activity, and a second step of reduction of the obtained 3 -hydroxypropionaldehyde with an enzyme having hydroxy aldehyde reductase activity, and genes enabling the microorganism to utilize sucrose as sole carbon source. The invention also concerns a new method for the biological preparation of 1,3-propanediol by fermentation, comprising cultivating said microorganism genetically modified, wherein the culture is performed in an appropriate medium comprising a source of sucrose, and recovering the 1,3 -propanediol being produced. In a preferred aspect of the invention, the source of sucrose is obtained from plant biomass.

The present disclosure provides recombinant bacteria with elevated 2-keto acid decarboxylase and alcohol transferase activities. Some recombinant bacteria further have elevated aldehyde dehydrogenase activity. Some recombinant bacteria further have reduced alcohol dehydrogenase and / or aldehyde reductase activity. Methods for the production of the recombinant bacteria, as well as for use thereof for production of various esters are also provided.