Search

Uracil-thymine dehydrogenase (UTDH), which catalyzes the irreversible oxidation of uracil to barbituric acid in oxidative pyrimidine metabolism, was purified from Rhodococcus erythropolis JCM 3132. The finding of unusual stabilizing conditions (pH 11, in the presence of NADP+ or NADPH) enabled the enzyme purification. The purified enzyme was a heteromer consisting of three different subunits. The enzyme catalyzed oxidation of uracil to barbituric acid with artificial electron acceptors such as methylene blue, phenazine methosulfate, benzoquinone, and α-naphthoquinone; however, NAD+, NADP+, flavin adenine dinucleotide, and flavin mononucleotide did not serve as electron acceptors. The enzyme acted not only on uracil and thymine but also on 5-halogen-substituted uracil and hydroxypyrimidine (pyrimidone), while dihydropyrimidine, which is an intermediate in reductive pyrimidine metabolism, and purine did not serve as substrates. The activity of UTDH was enhanced by cerium ions, and this activation was observed with all combinations of substrates and electron acceptors. • Uracil-thymine dehydrogenase was found to be a heteromer consisting of three different subunits. • Uracil-thymine dehydrogenase catalyzed oxidation of uracil with artificial electron acceptors. • The activity of uracil-thymine dehydrogenase was enhanced by cerium ions. [ABSTRACT FROM AUTHOR]

Mead acid (MA; 20:3ω9) is one of the ω9 series of polyunsaturated fatty acids (PUFAs). MA is used to inhibit the inflammation of joints and is applied to the medicinal or health food field. We aimed to construct MA-producing strains with disruption of the Δ12-desaturase gene (Δ1 2ds) via an efficient gene-targeting system using the lig4 -disrupted strain of Mortierella alpina 1S-4 as the host. The transformants showed a unique fatty acid composition that only comprised ω9-PUFAs and saturated fatty acids, while ω6-and ω3-PUFAs were not detected, and the total composition of ω9-PUFAs, including oleic acid (18:1ω9), 18:2ω9, 20:1ω9, 20:2ω9, and MA, was up to 68.4% of the total fatty acids. The MA production in the Δ1 2ds -disruptant reached 0.10 g/L (8.5%), which exceeded 0.050 g/L (4.6%) in the conventional Δ1 2ds -defective mutant JT-180. [ABSTRACT FROM AUTHOR]

To produce tomato juice with health-promoting functions, lactic acid bacteria (LAB) capable of converting l -glutamic acid in tomatoes into γ-aminobutyric acid (GABA) was screened from LAB stocks isolated from Japanese pickles. Lactiplantibacillus plantarum KB1253 was selected as the highest GABA producer among 74 strains of LAB stocks. gad gene expression and glutamic acid decarboxylation activity increased at low pH (3.0–3.5), whereas the growth decreased. Under optimal reaction conditions using resting cells as catalysts, this strain produced 245.8 ± 3.4 mM GABA. Furthermore, this strain produced 41.0 ± 1.1 mM GABA from l -glutamic acid in tomato juice under optimal fermentation conditions (pH 4.0, 20°Bx). This study may provide the basis for developing health-promoting functional foods rich in GABA from tomatoes and other agricultural products. [ABSTRACT FROM AUTHOR]

Enzymatic glycosylation is an industrially useful technique for improving the properties of compounds with hydroxy groups, and the biological activities of the resulting glycosides differ depending on the glycosylation position. Therefore, regioselective glycosyltransferases are required for precise synthesis of glycosides. We found that Rhizobium pusense JCM 16209T could catalyze the regioselective glycosylation of resveratrol. To identify the regioselective glycosyltransferase, two α-glucosidases of R. pusense JCM 16209T (RpG I and RpG II) were cloned and expressed in Escherichia coli. The molecular mass of purified recombinant RpG I and II was estimated to be 60 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). RpG I showed strong glycosylation activity toward resveratrol with 4′-selectivity of 98.3%. The enzyme activity was maximized at pH 8.0 and 50 °C, and enhanced in the presence of Cs+ and Li+ ions. The maximum molar yield of resveratrol 4′- O -α-glucoside from resveratrol reached 41.6% at 30 min, and the concentration of the product was 2.08 mmol L−1. Glycosylation activity was observed toward resveratrol as well as toward caffeic acid, ferulic acid, 6-gingerol, flavonoid, and isoflavonoid compounds with high regioselectivity, indicating that RpG I could glycosylate a wide range of substrates. To the best of our knowledge, there are few reports on microbial glycosyltransferases that are useful for regioselective glycosylation. This research could be the first step toward developing technologies for the precise synthesis of glycosides. [ABSTRACT FROM AUTHOR]

A wide variety of S-substituted cysteine derivatives occur in plant metabolites. For example, S -allyl- l -cysteine (SAC), mainly contained in garlic, gathers huge interest because of its favorable bioactivities for human health. However, conventional methods for preparing SAC suffer from several drawbacks with regard to efficiency and toxicity, which highlights the need for improved processes for SAC synthesis. This study aims to develop a novel bioprocess to produce SAC by microbial enzymes from easily available substrates. We found that Escherichia coli had the ability to synthesize SAC from allyl mercaptan, pyruvic acid, and ammonium sulfate. An enzyme purification through 3-step column chromatography, followed by determination of the N-terminal amino acid sequence revealed that tryptophanase (TnaA) was the enzyme responsible for SAC formation. Although the enzyme catalyzed the reversible reaction for synthesizing and degrading SAC, the degradation proceeded significantly faster than the synthesis. Interestingly, TnaA catalyzed the synthesis of a wide range of S-substituted cysteines with alkyl chains or aromatic rings, some of which are present in Allium and Petiveria plants. Our results showed a novel substrate specificity of TnaA toward various S-substituted cysteine. TnaA is a promising biocatalyst for developing a new process to supply various valuable S-substituted cysteine derivatives for medicinal and health-promoting applications. [ABSTRACT FROM AUTHOR]

Quantification of leuco-indigo is most important for Aizome , Japanese indigo-dyeing; however, there has been no convenient quantitative method. This study demonstrated that normal pulse voltammetry under quiescent conditions can be used to detect leuco-indigo. As a result of quantification of leuco-indigo in the depth direction in fermenting suspensions, the steady-state concentrations of leuco-indigo showed sigmoidal profiles in the depth direction. The steady state is caused by competitive reactions of microbial reduction of indigo and autoxidation of leuco-indigo by O 2 dissolved from the air interface of the suspension. In addition, we investigated the effects of stirring the suspension and adding some nutrients to the concentration profile. The weakened activity was partially recovered by the addition of ethanol and remarkably recovered by the addition of hipolypepton or glucose. Knowledge is essential for the proper management of indigo-dye-fermenting suspensions. [ABSTRACT FROM AUTHOR]

ω3-Docosapentaenoic acid (ω3-DPA), an ω3-polyunsaturated fatty acid (ω3-PUFA), is expected to have beneficial physiological functions to humans; however, because of its rarity in nature, it has not been fully analyzed. We isolated an ω3-DPA producing microorganism strain T7 from brackish areas in Japan. Although most oleaginous microorganisms rarely accumulate ω3-DPA (<5% of total lipid), strain T7 accumulated ω3-DPA with more than 20% of total fatty acids. The strain T7 was identified as a related species of Aurantiochytrium. In Aurantiochytrium sp. T7, ω3-DPA production reached 164 mg/L culture broth, and the ω3-DPA content reached 23.5% of the total fatty acids when cultivated in a medium containing 2% glucose as the carbon source and 1% yeast extract as the nitrogen source, with a salinity equivalent to 50% of that of seawater and a pH in the acidic range (pH < 5.5). Aurantiochytrium sp. T7 is a promising producer of high-purity ω3-DPA containing-lipid for the functional analysis of ω3-DPA whose physiological function has hardly been elucidated, and a useful strain for investigating the novel metabolic pathway of fatty acids. [ABSTRACT FROM AUTHOR]