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Properties of a recombinant β-glucosidase from polycentric anaerobic fungus Orpinomyces PC-2 and its application for cellulose hydrolysis

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Abstract

A β-glucosidase (BglA, EC 3.2.1.21) gene from the polycentric anaerobic fungus Orpinomyces PC-2 was cloned and sequenced. The enzyme containing 657 amino acid residues was homologous to certain animal, plant, and bacterial β-glucosidases but lacked significant similarity to those from aerobic fungi. Neither cellulose- nor protein-binding domains were found in BglA. When expressed in Saccharomyces cerevisiae, the enzyme was secreted in two forms with masses of about 110 kDa and also found in two forms associated with the yeast cells. K m and V max values of the secreted BglA were 0.762 mM and 8.20 µmol/(min·mg), respectively, with p-nitrophenyl-β-d-glucopyranoside (pNPG) as the substrate and 0.310 mM and 6.45 µmol/(min·mg), respectively, for the hydrolysis of cellobiose. Glucose competitively inhibited the hydrolysis of pNPG with a K i of 3.6 mM. β-Glucosidase significantly enhanced the conversion of cellulosic materials into glucose by Trichoderma reesei cellulase preparations, demonstrating its potential for use in biofuel and feedstock chemical production.

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Authors and Affiliations

  1. Fermentation Biotechnology Research Unit, National Center for Agricultural Utilization Research, USDA/ARS, 1815 N. University Street, 61604-3902, Peoria, IL

    Xin-Liang Li, Michael A. Cotta & Bruce S. Dien

  2. Department of Biochemistry & Molecular Biology and Center for Biological Resource Recovery, Life Science Building, University of Georgia, 30602, Athens, GA

    Lars G. Ljungdahl

  3. Laboratorio de Enzimlogia, Departamento de Biologia Celular, Universidade de Brasilia, 70910-900, Asa Norte, Brasilia-DF, Brazil

    Eduardo A. Ximenes & Carlos R. Felix

  4. Division of Microbiology, National Center for Toxicological Research, US Food and Drug Administration, 3900 NCTR Road, 72079-9501, Jefferson, AR

    Huizhong Chen

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  1. Xin-Liang Li
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  2. Lars G. Ljungdahl
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  3. Eduardo A. Ximenes
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  4. Huizhong Chen
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Correspondence to Xin-Liang Li.

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Names are necessary to report factually on available data; however, the USDA neither guarantees nor warrants the standard of the product, and the use of the name by USDA implies no approval of the product to the exclusion of others that may be suitable.

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Li, XL., Ljungdahl, L.G., Ximenes, E.A. et al. Properties of a recombinant β-glucosidase from polycentric anaerobic fungus Orpinomyces PC-2 and its application for cellulose hydrolysis. Appl Biochem Biotechnol 113, 233–250 (2004). https://doi.org/10.1385/ABAB:113:1-3:233

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  • DOI: https://doi.org/10.1385/ABAB:113:1-3:233

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