A β‐glucosidase gene (bgl3) from Streptomyces sp. strain QM‐B814: Molecular cloning, nucleotide sequence, purification and characterization of the encoded enzyme, a new member of family 1 glycosyl hydrolases

Josep A. PEREZ‐PONS, Antonio CAYETANO, Xavier REBORDOSA, Jorge LLOBERAS, Alicia GUASCH, Enrique QUEROL

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Abstract

A β‐glucosidase gene (bgl3) from Streptomyces sp. QM‐B814 (American Type Culture Collection 11238) has been cloned by functional complementation of a β‐glucosidase‐negative mutant of Streptomyces lividans. An open‐reading frame of 1440 nucleotides encoding a polypeptide of 479 amino acids was found by sequencing. The encoded protein (Bgl3) shows extensive similarity (over 45% identity) with β‐glycosidases from family‐1 glycosyl hydrolases. The cloned enzyme, purified following ammonium sulphate precipitation and two chromatographic steps, is monomeric with molecular mass 52.6 kDa, as determined by mass spectrometry, and an isoelectric point of pI 4.4. The enzyme appears to be a β‐glucosidase with broad substrate specificity, is active on cellooligomers, and performs transglycosylation reactions. The estimated apparent Km values for p‐nitrophenyl‐β‐d‐glucopyranoside and cellobiose are 0.27 mM and 7.9 mM, respectively. The Ki values for glucose and δ‐gluconolactone, using p‐nitrophenyl‐β‐d‐glucopyranoside as a substrate, are 65 mM and 0.08 mM, respectively. The purified enzyme has a pH optimum of pH 6.5 and the temperature optimum for activity is 50°C. Copyright © 1994, Wiley Blackwell. All rights reserved
Original languageEnglish
Pages (from-to)557-565
JournalEuropean Journal of Biochemistry
Volume223
Issue number2
DOIs
Publication statusPublished - 1 Jan 1994

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