Process Biotechnology

Microbial Biotechnology

Electronic Journal of Biotechnology ISSN: 0717-3458 Vol. 10 No. 2, Issue of January 15, 2007
© 2007 by Pontificia Universidad Católica de Valparaíso -- Chile Received April 5, 2006 / Accepted October 26, 2006
DOI: 10.2225/vol10-issue2-fulltext-4
RESEARCH ARTICLE

Purification and characterization of β-glucosidase from Melanocarpus sp. MTCC 3922

Jatinder Kaur
Department of Microbiology
Guru Nanak Dev University
Amritsar 143005, India
Tel: 91 183 2258506
E-mail: jatinderkaur77@yahoo.com 

Bhupinder S. Chadha*
Department of Microbiology
Guru Nanak Dev University
Amritsar 143005, India
Tel: 91 183 2258802
Fax: 91 183 2258820
E-mail: chadhabs@yahoo.com 

Badhan A. Kumar
Department of Microbiology
Guru Nanak Dev University
Amritsar 143005, India
Tel: 91 183 2258802 09. Ext. 3317
E-mail: badhanak@yahoo.co.in

Ghatora, S. Kaur
Department of Microbiology
Guru Nanak Dev University
Amritsar 143005, India
Tel: 91 1852 228868
E-mail: ghatorask@yahoo.com 

Harvinder S. Saini
Department of Microbiology
Guru Nanak Dev University
Amritsar 143005, India
Tel: 91 183 2258802 09. Ext. 3318
E-mail: sainihs@yahoo.com

*Corresponding author

Financial support: The financial support in form of project fellowship provided by Guru Nanak Dev University, Amritsar (India).

Keywords: β-glucosidase, Melanocarpus sp., purification, substrate specificity, transglycosylation.

Abbreviations:

IEF: Isoelectric focusing
MTCC: Microbial Type Culture Collection
PBE: Poly-buffer exchanger
pNPG: p-nitrophenyl-β-D-glicopyronoside
SSF: Solid-state fermentation
YpSS: Yeast potato soluble starch agar
Abstract
Full Text

This study reports the purification and characterization of β-glucosidase from a newly isolated thermophilic fungus, Melanocarpus sp. Microbial Type Culture Collection (MTCC) 3922. The molecular weight of β-glucosidase was determined to be ~ 92 and 102 kDa with SDS PAGE and gel filtration, respectively, and pI of ~ 4.1. It was optimally active at 60ºC and pH 6.0, though was stable at 50ºC and pH 5.0 - 6.0. The presence of DTT, mercaptoethanol andmetal ions such as Na+, K+, Ca2+, Mg2+and Zn2+ positively influenced the activity of β-glucosidase but the activity was inhibited in the presence of CuSO4. β-Glucosidase recognized pNP- β-glucopyranoside (pNPG) as the preferred substrate, and showed very low affinity for pNP- β-D-cellobioside. Km and Vmax for the hydrolysis of pNPG by β-glucosidase was calculated as 3.3 mM and 43.68 :molmin-1mg protein-1, respectively and kcat was quantified as 4 x 103 min-1. β-Glucosidase activity was enhanced appreciably in the presence of alcohols (methanol and ethanol) moreover, purified β-glucosidase showed putative transglycosylation activity that was positively catalyzed in presence of methanol as an acceptor molecule.
Supported by UNESCO / MIRCEN network