Yamada et al. found a novel aldehyde reductase in a red yeast, Sporobolomyces salmonicolor (35-37), and succeeded in cloning the aldehyde reductase gene from this strain (38).
Aldehyde reductase was isolated in a crystalline form from cells of S. salmonicolor and was characterized in some detail (35-37). It should be noted that the cellular content of the enzyme comprised more than 4.5% of the total ex-tractable protein. The enzyme has a monomeric form of about 35 kDa. The enzyme absolutely requires NADPH as a cofactor. The substrate specificity of the enzyme is shown in Table 1. The enzyme did not catalyze the reduction of any keto acids or monoketones tested. Several aldehydes such as p-nitrobenzaldehyde, p-chlorobenzaldehyde, pyridine-3-aldehyde, or D-glyceraldehyde were readily reduced. These aldehydes are typical substrates for aldo-keto reductase family members, as already described. Qui-nones, such as p-benzoquinone, a-naphthoquinone, /-naphthoquinone, and menadione, which are good substrates for carbonyl reductases (14), were not reduced. Although no detectable oxidation of NADPH was observed with D-glucuronate, D-glucose, D-xylose, or D-galactose as the substrate at a low concentration (0.2 mM), the enzyme effectively catalyzed the reduction of these compounds when they were present at a high concentration (100 mM). The Km values for these substrates are quite high in comparison with those for other substrates, which suggests that the affinity of the enzyme for these substrates is very low. Furthermore, the enzyme effectively catalyzed the reduction of 4-haloacetoacetate esters to the corresponding (R)-4-halo-3-hydroxybutanoate esters (100% ee) (35,36). Because the reduction products derived from 4-haloaceto-acetate esters are optically active alcohols, the enzyme can recognize the stereopositions of these substrates.
2-Chloroacetoacetate esters were also effective substrates. Initial velocity and inhibition studies indicated that the enzyme reaction sequence is ordered, with NADPH binding to the free enzyme, and NADP+ being the last product to be released. Quercetin, dicoumarol, and some SH-reagents inhibited the enzyme activity. These results ofen-zymological studies suggested that the enzyme of S. sal-monicolor belongs to the aldo-keto reductase superfamily.
cDNA coding the aldehyde reductase was cloned from cells of S. salmonicolor and sequenced (38). The aldehyde reductase gene comprises 969 bp and encodes a polypeptide of 35,232 Da. The deduced amino acid sequence shows a high degree of similarity to that of other members of the aldo-keto reductase superfamily. The identities between S. salmonicolor aldehyde reductase and the bovine (39), rat (40), human (7), rabbit (41), and barley (42) aldose reductases, human aldehyde reductase (7), and xylose reductase from Pichia stipitis (43) are 45.1, 45.0, 43.7, 42.9, 39.5, 43.9, and 37.7%, respectively. In general, aldo-keto reduc-tases exhibit greater similarity in their N-terminal regions than in their C-terminal regions (Fig. 1). Analysis of the genomic DNA sequence indicated that the aldehyde reduc-tase gene was interrupted by six introns (two in the 5' non-coding region and four in the coding region).
Table 1. Substrate Specificity of the Aldehyde Reductase from S. salmonicolor
Relative activity (%)
Relative activity (%)
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