title: Purification and Characterization of Isoamyl Alcohol Oxidase ("Mureka"-Forming Enzyme).
abstract: Isoamyl alcohol oxidase (IAAOD) was purified to apparent homogeneity on SDS-PAGE from ultrafiltration (UF) concentrated sake. IAAOD was a glycoprotein, a monomeric protein with an apparent molecular mass of 73 and 87 kDa, by SDS-PAGE and gel filtration on HPLC, respectively. IAAOD showed high substrate specificity toward C5 branched-chain alkyl alcohol (isoamyl alcohol), and no activity toward shorter (C1-C4) or longer (C7-C10) alkyl alcohols tested. IAAOD was stable between pH 3.0-6.0 at 25°C. The optimum pH was 4.5 at 35°C. Heavy metal ions, p-chloromercuribenzoate (PCMB), hydrazine, and hydroxylamine strongly inhibited the enzyme activity, and an anti-oxidant like L-ascorbate did also. Isovaleraldehyde was produced markedly in pasteurized sake by adding purified IAAOD, therefore, we concluded that it was the enzyme that causes formation of mureka, an off-flavor of sake, the main component of which is isovaleraldehyde.
authors.affiliation: a Research and Development Department, Hakutsuru Sake Brewing Co. Ltd.
authors.forename: N
authors.initials: N
authors.lastname: Yamashita
authors.affiliation: a Research and Development Department, Hakutsuru Sake Brewing Co. Ltd.
authors.forename: T
authors.initials: T
authors.lastname: Motoyoshi
authors.affiliation: a Research and Development Department, Hakutsuru Sake Brewing Co. Ltd.
authors.forename: A
authors.initials: A
authors.lastname: Nishimura
keywords: Aspergillus oryzae; alcohol oxidase; mureka; off-flavor enzyme