The present invention relates to a process for producing efficiently glutamic acid derivatives (including salts thereof) such as monatin by converting a substituted α-keto acid of formula (1) into a glutamic acid derivative of formula (2) in the presence of an enzyme catalyzing conversion of the same.

Process for producing glutamate derivatives

By simultaneously carrying out an isomerization reaction at position 2 of monatin in different configurations at positions 2 and 4 in the presence of an aldehyde under a condition of pH 4 to 11 in a mixture solvent of water and an organic solvent, and the crystallization of monatin in the same configurations at positions 2 and 4 or a salt thereof, monatin useful as a sweetener, particularly optically active monatin can efficiently be produced.

Method for producing monatin

Indole pyruvic acid (IPA) is a versatile platform intermediate and building block for a number of high-value products in the pharmaceutical and food industries. It also has a wide range of applications, such as drugs for the nervous system, cosmetics, and luminophores. Chemical synthesis of IPA is a complicated and costly process. Moreover, through the biosynthesis route employing l-amino acid oxidase, the byproduct hydrogen peroxide leads the degradation of IPA. TdiD, identified as a specific tryptophan aminotransferase, could be an alternative solution for efficient IPA biosynthesis. Escherichia coli strain W3110, which demonstrates basic production when supplied with tryptophan, was engineered for IPA biosynthesis. Several strategies were implemented to improve IPA production. First, through incorporating the codon-optimized tdiD into W3110, IPA levels increased from 41.54 ± 1.26 to 52.54 ± 2.08 mg/L. Second, after verifying the benefit of an increased phenylpyruvate pool, a YL03 strain was constructed based on a previously reported mutant strain of W3110 with a plasmid carrying aroF
 fbr
 and pheA
 fbr
 to further improve IPA production. The recombinant YL03 strain accumulated IPA at 158.85 ± 5.36 mg/L, which was 3.82-fold higher than that of the wild-type W3110 strain. Third, optimization of tdiD
 co
 expression was carried out by replacing the Trc promoter with a series of constitutively active promoters along with increasing the plasmid copy numbers. The highest IPA production was observed in YL08, which achieved 236.42 ± 17.66 mg/L and represented a greater than 5-fold increase as compared to W3110. Finally, the effects of deletion and overexpression of tnaA on IPA biosynthesis were evaluated. The removal of tnaA led to slightly reduced IPA levels, whereas the overexpression of tnaA resulted in a considerable decline in production. This study illustrates the feasibility of IPA biosynthesis in E. coli through tdiD. An efficient IPA producing strain, YL08, was developed, which provides a new possibility for biosynthesis of IPA. Although the final production was limited, this study demonstrates a convenient method of IPA synthesis.

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Metabolic engineering of indole pyruvic acid biosynthesis in Escherichia coli with tdiD.

The present invention provides a methodology for improving a yield of 2R,4R-Monatin. Specifically, the present invention provides a method for producing 2S,4R-Monatin or a salt thereof, comprising contacting 4R-IHOG with an L-amino acid aminotransferase in the presence of an L-amino acid to form the 2S,4R-Monatin; a method for producing 2R,4R-Monatin or a salt thereof, comprising isomerizing the 2S,4R-Monatin to form the 2R,4R-Monatin; and the like. These production methods may further comprise condensing indole-3-pyruvate and pyruvate to form the 4R-IHOG, and deaminating a tryptophan to form the indole-3-pyruvate.

Method for producing monatin using an l-amino acid aminotransferase

A method for producing benzaldehyde is provided. Benzaldehyde is produced from L-phenylalanine or a carbon source by using microorganism(s) having amino acid deaminase, 4-hydroxymandelate synthase, (S)-mandelate dehydrogenase, and benzoylformate decarboxylase.

Method for producing benzaldehyde

It is an object of the present invention to provide a procedure for realizing inexpensive and simple production of 3-indole-pyruvic acid. A transformant is made using a polynucleotide having a specific nucleotide sequence encoding a protein having an oxidase activity, and oxidase is generated by culturing the transformant in a medium to accumulate the oxidase in the medium and/or the transformant. Further, tryptophan is converted into 3-indole-pyruvic acid in the presence of the transformant and/or a culture thereof to produce 3-indole-pyruvic acid.

Novel oxidase gene and method for producing 3-indole-pyruvic acid by utilizing the gene

PROBLEM TO BE SOLVED: To provide a method for efficiently producing D-arabitol, and a method for producing D-xylulose and xylitol using the method. SOLUTION: The method for producing xylitol comprises the steps of: producing D-arabitol in a medium by culturing a microorganism which belongs to the genus Metschnikowia, and can produce D-arabitol (e.g. M pulcherrima, M reukaufii, M bicuspidata, M. lutana, and M. zobellii) aerobically in a medium containing saccharide(s) as the main carbon source(s) to form D-arabitol in the medium, inoculating/culturing a microorganism which has an ability of producing D-xylulose from D-arabitol to form D-xylulose in the broth, and inoculating/culturing a microorganism which has an ability of transforming D-xylulose to xylitol to the broth containing D-xylulose to collect xylitol formed.

Production of d-arabitol, d-xylulose and xylitol

It is intended to provide a means of achieving the economical and convenient production of 3-indole-pyruvic acid By using a polynucleotide that has a definite base sequence encoding a protein having an oxidase activity, a transformant is constructed Then, this transformant is cultured in a medium and thus oxidase is produced by accumulating the oxidase in the medium and/or the transformant Further, tryptophan is converted into 3-indole-pyruvic acid in the presence of the above-described transformant and/or the culture thereof to thereby produce 3-indole-pyruvic acid

Novel oxidase gene and method of producing 3-indole-pyruvic acid using the gene

PURPOSE: To provide a method for efficiently producing D-arabitol, and a method for producing D-xylulose and xylitol using the method. CONSTITUTION: The method for producing xylitol comprises the steps of: producing D-arabitol in a medium by culturing a microorganism which belongs to the genus Metschnikowia, and can produce D-arabitol (e.g. M pulcherrima, M reukaufii, M bicuspidata, M. lutana, and M. zobellii) aerobically in a medium containing saccharide(s) as the main carbon source(s) to form D-arabitol in the medium, inoculating/culturing a microorganism which has an ability of producing D-xylulose from D-arabitol to form D-xylulose in the broth, and inoculating/culturing a microorganism which has an ability of transforming D-xylulose to xylitol to the broth containing D-xylulose to collect xylitol formed.

Suzuki Shunichi

Papers

Novel oxidase gene and method for producing 3-indole-pyruvic acid by utilizing the gene

Production of d-arabitol, d-xylulose and xylitol

Novel oxidase gene and method of producing 3-indole-pyruvic acid using the gene

- - Methods for producing -arabitol -xylulose and xylitol

Methods for making 3-indole-pyruvic acid from tryptophan using a tryptophan deaminase