Teruzo Miyoshi

Characterization of aldehyde reductase of Sporobolomyces salmonicolor

An NADPH-dependent aldehyde reductase (EC 1.1.1.2) isolated from Sporobolomyces salmonicolor AKU 4429 was further characterized. The enzyme also catalyzed the reductions of D-glucuronate, D-glucose, D-xylose and D-galactose at high concentrations. Km values for D-glucuronate and D-glucose are 345 and 4270 mM, respectively. Quercetin, dicoumarol and some SH-reagents inhibited the enzyme activity. NH2-terminal amino acid sequence analysis showed that the S. salmonicolor enzyme is partially the same as the aldo-keto reductase family proteins in primary protein structure.

Purification and characterization of 2-alkyne-1-ol dehydrogenase induced by 2-butyne-1,4-diol in Fusarium merismoides B11

Abstract 2-Alkyne-1-ol dehydrogenase from the mycelia of Fusarium merismoides B11 was purified 320-fold by fractionation with ammonium sulfate followed by chromatographies on O-(diethylaminoethyl)-cellulose, calcium phosphate gel and Sephadex G-200. On ultracentrifugation, the enzyme gave a single, sharp, symmetrical schlieren peak with a sedimentation coefficient (S20,w) of 22.3 S. Activity is dependent on both NAD+ and NADP+, especially the former. The enzyme has an optimal pH of 8.2, and Km value of 9.1·10−3 M for 2-butyne-1,4-diol. The general formula for a well-oxidizable substrate is RCCCH2OH, where R may be H, CH3, CH2OH and CCCH2OH with the exception of 1,4-butanediol. The activity is sensitive to thiol reagents but not to metal-chelating agents. The enzyme is only formed when 2-butyne-1,4-diol was used as the sole source of carbon.

D-Threonine Aldolase and Its Application to D -β-Hydroxy-£-Amino Acid Synthesis

D - β -Hydroxy - α - amino acids such as D-threonine are useful as the intermediate in the synthesis of chiral compounds such as antibiotics. It is difficult to synthesize D-threonine stereospecifically because of having two asymmetric carbons at α - and β -positions. We have found two types of threonine aldolase specific for other stereoisomers than L-threonine. D-Threonine aldolase (DTA) catalyzes the reversible aldol cleavage of D-threonine into glycine and acetaldehyde [1]. This enzyme can provide the synthetic route of D-threonine via the reverse synthesis. In this report, we will describe the isolation and characterization of DTA, and also describe the method of producing D-threonine stereospecifically from glycine and acetaldehyde. This method is based on the phenomena that glycine and threonine participate in the complex formation with metal ions and the enzymatic reaction catalyzed by DTA, as might be referred to as the “metal complex-assisted enzyme reaction”.