99 results about "Glutamate Dehydrogenase (NADP+)" patented technology

The invention is about a method of measuring the content of blood ammonia, and it also concerns the reagent box of blood ammonia diagnosis. This invention belongs to the field of medical testing and measuring technology. The reagent box is consisted of buffer solution, 2ú¡ketoglutarate, reduced coenzyme, adenosine triphosphate, phosphoenolpyruvate, glutamate dehydrogenase, glutamate kinase, pyruvate kinase, lactate dehydrogenase and stabilizer. Firstly, we cause an enzyme-coupled reaction through mixing the sample and the reagent according to a certain proportion of volume; secondly, put the final reactant under the biochemical analyzer and test the absorbance variational situation (speed) of dominant wavelength; then we can get the content of blood ammonia. By using this invention, we can get the necessary measuring result with high sensitiveness and fine precision through biochemical analyzer, and the result would not be contaminated by material of internal and exogenous sources. Thus, this method can be conveniently promoted and applied.

A genetically engineered bacterium producing 2-phenylethanol and an applying method thereof are disclosed. The recombinant bacterium is constructed by coexpressing aromatic amino acid aminotransferase and exogenous phenylpyruvate decarboxylase (Aro10) (or indole-3-pyruvic acid decarboxylase), phenylethanol dehydrogenase or phenylacetaldehyde reductase in escherichia coli to obtain the recombinant bacterium producing the 2-phenylethanol. Glutamate dehydrogenase is coexpressed on this basis to achieve self-balancing cofactor circulation, thus increasing the yield of the 2-phenylethanol. The conversion ratio of the 2-phenylethanol prepared through the recombinant bacterium is high. The bacterium and the method have important application value in production of the 2-phenylethanol in the future.

A non-naturally occurring microbial organism has at least one exogenous nucleic acid encoding a MEK pathway enzyme expressed in a sufficient amount to produce MEK. The MEK pathway includes an enzyme selected from an acetoacetyl-CoA dehydrogenase (bifunctional), an acetoacetyl-CoA aldehyde dehydrogenase, a 3-oxobutyraldehyde reductase, a 3-oxobutanol dehydratase, an MEK oxidoreductase, a 3-oxobutyraldehyde aminotransferase, a 4-aminobutan-2-one deaminase, a 2-amino-4-ketopentanoate (AKP) thiolase, an AKP aminotransferase, a 2,4-dioxopentanoate decarboxylase, an AKP deaminase, an acetylacrylate decarboxylase, an AKP decarboxylase, a glutamate dehydrogenase, a 3-oxobutyraldehyde oxidoreductase (aminating) and an AKP oxidoreductase (aminating). A 2-butanol pathway further includes an MEK reductase. A method for producing MEK or 2-butanol includes culturing these organisms under conditions and for a sufficient period of time to produce MEK or 2-butanol.

The invention discloses a new strain MP688 of Methylovorus sp., of which the preservation number is CGMCC No.4096, the preservation date is August 20th, 2010, and the preservation unit is China General Microbiological Culture Collection Center. The strain MP688 is separated from soil by using a GDH (glutamate dehydrogenase) recombinase method of PQQ (pyrroloquinoline quinone). By using the Methylovorus sp. strain disclosed by the invention, the yield of PQQ can reach 125 mg/L in a conventional culture medium under conventional culture conditions; and the Methylovorus sp. strain can be used for industrial production of PQQ, and has important meaning for promoting the industrialization of PQQ.

The present invention relates generally to the field of molecular biology and concerns a method for improving various plant growth characteristics by modulating expression in a plant of a nucleic acid encoding a GDH (Glutamate DeHydrogenase) polypeptide. The present invention also concerns plants having modulated expression of a nucleic acid encoding a GDH polypeptide, which plants have improved growth characteristics relative to corresponding wild type plants or other control plants. The invention also provides constructs useful in the methods of the invention. The present invention relates generally to the field of molecular biology and concerns a method for enhancing various economically important yield-related traits in plants. More specifically, the present invention concerns a method for enhancing yield-related traits in plants by modulating expression in a plant of a nucleic acid encoding a FLA-like (Fasciclin-like) polypeptide. The present invention also concerns plants having modulated expression of a nucleic acid encoding a FLA-like polypeptide, which plants have enhanced yield-related traits relative to control plants. The invention also provides constructs comprising FLA-like- encoding nucleic acids, useful in performing the methods of the invention. The present invention relates generally to the field of molecular biology and concerns a method for enhancing yield-related traits in plants by modulating expression in a plant of a nucleic acid encoding a SAUR polypeptide. The present invention also concerns plants having modulated expression of a nucleic acid encoding a SAUR polypeptide, which plants have enhanced yield-related traits relative to corresponding wild type plants or other control plants. The invention also provides constructs useful in the methods of the invention. Furthermore, the present invention also relates to a SAUR-based protein complex. It further relates to the use of the complex to enhance yield-related traits, and to a method for stimulating the complex formation, by overexpressing at least two members of the complex. The present invention relates generally to the field of molecular biology and concerns a method for enhancing yield traits in plants by modulating expression in a plant of a nucleic acid encoding a dehydroascorbate reductase (DHAR) polypeptide. The present invention also concerns plants having modulated expression of a nucleic acid encoding a DHAR polypeptide, which plants have enhancing yield traits relative to corresponding wild type plants or other control plants. The invention also provides constructs useful in the methods of the invention.

The invention discloses an enzyme combination for producing L-phosphinothricin. The enzyme combination comprises glutamate dehydrogenase and a coenzyme regenerating enzyme, wherein the coenzyme regenerating enzyme is alcohol dehydrogenase, formate dehydrogenase and phosphite dehydrogenase. The invention also discloses a production method of L-phosphinothricin, 4-(methylhydroxyphosphinyl)-2-oxobutyric acid is used as a raw material, NH<4><+>, a coenzyme NADP<+>/NADP, and a coenzyme regeneration substrate are added, and then the enzyme combination is used for catalysis, wherein the glutamate dehydrogenase is used to catalyze a reaction of 4-(methylhydroxyphosphinyl)-2-oxobutyric acid to obtain L-phosphinothricin, and the coenzyme regenerating enzyme is used to reduce NADP<+> to NADP. According to the enzyme combination and the production method of L-phosphinothricin provided by the invention, by-products produced are very easy to remove, a post-treatment process of the product is simplified, the total yield of the product is greater than 95%, and the production cost of L-phosphinothricin is reduced, so that the method is a green, environment-friendly, and low-carbon process route, and is suitable for large-scale industrial production applications.

The invention relates to a kit for diagnosing/measuring glycine by utilizing the technologies of the enzymatic doubling amplification method, the enzymic colorimetry and the enzyme linked immunosorbent assay. The invention further relates to a method, a principle and the composition and the components of a reagent for measuring the concentration of the glycine, and belongs to the technical field of medical/food inspection and measurement. The main components of the kit include a buffer solution, coenzyme, a 2-ketoglutaric acid, guanosine monophosphate, glycine aminotransferase, glutamate dehydrogenase, guanosine monophosphate reduced enzyme and a stabilizer. Through mixing a sample and the reagent by a certain volume ratio, a series of enzymatic reactions occur, then the reactant is placed under an ultraviolet/visible light analyzer, and the degree/velocity of the increase in absorbance at 340 nm of the dominant wavelength is detected, thereby measuring the active concentration of the glycine.

The invention relates to the technical field of urea detection, in particular to a preparation method of a urea determination reagent ball, the reagent ball and a detection chip. The embodiment of theinvention provides a preparation method of a urea determination reagent ball. The method comprises the following steps of mixing a certain dosage of buffer solution, disodium ethylene diamine tetraacetate, alpha-ketoglutarate, glutamate dehydrogenase, urease, reduced nicotinamide adenine dinucleotide and excipient with water to form a mixed solution, dropwise adding the mixed solution into liquidnitrogen to enable liquid drops of the mixed solution to form ice balls, and freeze-drying the ice balls to obtain spherical urea determination reagent balls. The urea determination reagent ball prepared by the method has good thermal stability, can be stored for 8 days at a high temperature of 37 DEG C, is not easy to deteriorate in the storage process, has a wide effective test range on urea inblood, and can test the highest concentration of blood urea as high as 35mmol/L.

Provided herein are SIRT4 compositions and methods of use thereof. The invention provides functional information for use in the identification and design of compounds that modulate SIRT4 enzyme activity (e.g., inhibition of fatty acid oxidation, ADP ribosylation, and/or downregulation of glutamate dehydrogenase), and to the compounds identified by such methods and the research, diagnostic and therapeutic uses of such compounds.

The present invention relates to glutamate dehydrogenase mutants and their application in preparation of L-phosphinothricin. The amino acid sequences of the glutamate dehydrogenase mutants are as shown in SEQ ID NO. 1˜9, 11, 13, 15, 17˜19 and 22. By means of molecular engineering, mutating the specific alanine in glutamate dehydrogenase substrate-binding pocket into glycine and/or mutating the specific valine in glutamate dehydrogenase substrate-binding pocket into alanine, the present invention has obtained NADPH-specific glutamate dehydrogenase mutants with high enzyme activity in catalyzing the substrate 2-oxo-4-[(hydroxy)(methyl)phosphinoyl]butyric acid or its salt for L-phosphinothricin preparation or NADH-specific glutamate dehydrogenase mutants with catalytic activity toward PPO; this has significantly improved substrate conversion, and increased the product concentration of the L-phosphinothricin preparation process.

The invention relates to a mutant gene and coding protein of glutamate dehydrogenase of Bacillus natto, belonging to the field of biotechnology. The base sequence of the mutant gene of the coding gene of glutamate dehydrogenase of Bacillus natto is shown as follows: SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7, SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13 or SEQ ID NO: 15; the protein amino acid sequence of the mutant gene of the coding gene of glutamate dehydrogenase of Bacillus natto is shown as follows: SEQ ID NO: 4, SEQ ID NO: 6, SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14 or SEQ ID NO: 16. The invention also relates to a recombinant vector containing the mutant gene. When the mutation of the invention is introduced to wild strains by homologous recombination, gene engineering bacteria with low ammonia yield and unchanged growth speed is obtained, thus producing natto without ammonia odor.

The invention belongs to the field of plant genetic engineering, and discloses clone and application of a fungus PcGDH (Pleurotuscystidiosus Glutamate Dehydrogenase) gene for improving efficient utilization of nitrogen. The NADP (H) (Nicotinamide Adenine Dinucleotide Phosphate) enzymatic activity of a PcGDH in-vitro positive reaction is larger than that of a reverse reaction, that is the PcGDH protein is inclined to utilize NH4+ to translate alpha-oxoglutarate into glutamic acid. In addition, the affinity of the PcGDH to NH4+ is larger than glutamic acid. By virtue of a genetic engineering technology, the PcGDH gene is over-expressed in rice in a heterogeneous manner to improve utilization of nitrogen by transgenic rice, improve growth of the transgenic rice and increase absolute plant and thousand seed weight of the transgenic rice. Therefore, the PcGDH gene for improving the rice nitrogen can be utilized to culture novel rice products with good agronomic characters.

The invention discloses a gene mining method combining a functional sequence and structural simulation, an NADH preference type glufosinate-ammonium dehydrogenase mutant and an application. The gene mining method comprises the following steps: (1) analyzing a characteristic sequence of NADH type glutamate dehydrogenase; (2) searching a gene pool according to the characteristic sequence; (3) performing clustering analysis and protein structure simulation on the searched genes; and (4) selecting a gene with high gene polymerization degree and a protein structure similar to that of known glufosinate-ammonium dehydrogenase as a candidate gene. The method comprises the following steps: carrying out gene mining to obtain wild glufosinate-ammonium dehydrogenase which is derived from Lysinibacillus composti and has an amino acid sequence shown as SEQ ID No.2, carrying out mutation screening to obtain an NADH preference type glufosinate-ammonium dehydrogenase mutant, and selecting a mutation site from one of the following: (1) A144G-V375F-M91A; (2) A144G-V345A-M91A; and (3) A144G. The mutant enzyme can be subjected to catalytic reaction by using cheap coenzyme NAD.

The invention discloses an L-glutamate dehydrogenase mutant. The sequence of the L-glutamate dehydrogenase mutant is obtained by mutating a 175 amino acid residue A of the sequence shown in SEQ IDNO. 1 into G and mutating a 386 amino acid residue V into an amino acid residue with smaller steric hindrance. The invention also discloses application of the L-glutamate dehydrogenase mutant inpreparation of L glufosinate-ammonium or salts thereof. When the L-glutamate dehydrogenase mutant is used for preparing the L-glufosinate-ammonium or salts thereof, compared with the L-glutamate dehydrogenase mutant which is only mutated at the 175 site or 386 site, the L-glutamate dehydrogenase mutant has higher enzyme activity, thereby improving the action efficiency of the enzyme, decreasing the reaction cost and being beneficial to industrial production.

The invention discloses a kit for determining homocysteine and a preparation method thereof. The kit is composed of dual-liquid components of a reagent R1and a reagent R2 which are independent from each other. The reagent R1 is prepared from a phosphate buffer, sodium chloride, S-adenosylmethionine, a reduced coenzyme II, tri(2-carboxyl) phosphine hydrogen chloride and sodium azide; the reagent R2 is prepared from phosphate buffer, HCY transmethylase, glutamate dehydrogenase, S-adenosylhomocysteine hydrolase, bovine serum albumin and sodium azide. The preparation method comprises the steps that the reagents are prepared according to the component content; a to-be-tested sample is mixed with the reagent R1 and the reagent R2 to perform a sufficient reaction; a fully automatic biochemical analyzer is used for determining the reacted absorptivity difference; according to the absorptivity change value, the concentration of homocysteine in the sample is worked out. The kit has the advantages of being convenient to operate, high in accuracy and the like.

The invention discloses a glutamate dehydrogenase mutant and application thereof, and belongs to the technical field of enzyme engineering and microbial engineering. By adopting the method provided bythe invention, a brand-new method for synthesizing (R) -4-aminovaleric acid by a biological method is provided; wild glutamate dehydrogenase has no measurable catalytic activity, but the glutamate dehydrogenase mutant has catalytic activity, and the catalytic activity of the mutants K116S/N348L, K116E/N348M and K116Q/N348M can reach 1.87 U/mg, 3.16 U/mg and 4.55 U/mg respectively, so that the catalytic activity of the 4-oxovaleric acid is improved from zero. The biological method for synthesizing (R) -4-aminovaleric acid provided by the invention is green, environment-friendly and efficient,and has a wide industrial application prospect.

The invention provides a urea detection kit as well as a preparation method and a use method thereof. The kit comprises a reagent R1 and a reagent R2, wherein the reagent R1 comprises a Tris buffer solution, adenosine diphosphate, urease and glutamate dehydrogenase, and the reagent R2 comprises a Tris buffer solution, reduced coenzyme I and alpha-ketoglutaric acid. The kit provided by the invention is simple in component, maintains the advantages of higher sensitivity, wide linear range, low cost, strong anti-interference capability and high accuracy by adopting an enzyme coupling rate methodin an enzyme method, and can be universally applied to full-automatic and semi-automatic analyzers.