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KMID : 0545119910010030188
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Journal of Microbiology and Biotechnology
1991 Volume.1 No. 3 p.188 ~ p.196
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Purification and Characterization of β-Glucosidase from Penicillium verruculosum
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Chun, Soon Bai
Kim, Dong Ho/Kim, Kang Hwa/Chung, Ki Chul
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Abstract
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The β-glucosidase was purified to homogeneity from the culture filtrate of P. verruculosum by column chromatography. The enzyme was a glycoprotein with a relative size of approximately 220 kDa with an isoelectric point of 4.8, which was composed of dimeric protein of 105 kDa. The enzyme was stable up to 60℃ and the presence of glycerol significantly increased its thermostability. The enzyme was found to hydrolyze both β-aryl and β-alkyl-glucosides in addition to β-glucosyl glucose and catalyzed glucosyl transfer to cellobiose. The enzyme attacked laminarin in an exotype-like fashion. The apparent Km´s of the enzyme toward cellobiose, laminaribiose, laminarin were 0.53mM, 0.35mM and 1.11mM, respectively. Glucose and glucono-δ-lactone were competitive inhibitors for the enzyme. Copper (Cu^2+), mercury (Hg^2+) and p-chloromercuribenzoate were strong inhibitors of the enzyme. The immunoblotting result revealed that one form of β-glucosidase was biosynthesized, irrespective of carbon sources used. Polyacrylamide gel electrophoresis analysis of the in vitro translated product of total RNA from avicel grown mycelium established that the P. verruculosum β-glucosidase precursor was approximately 95 kDa in size. The amino acid composition and N-terminal amino acid sequence are given.
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