Purification and characterization of a novel ?-galactosidase from Bacillus sp MTCC 3088

Chakraborti, S and Sani, R K and Banerjee, U C and Sobti, R C (2000) Purification and characterization of a novel ?-galactosidase from Bacillus sp MTCC 3088. Journal of Industrial Microbiology and Biotechnology, 24 (1). pp. 58-63. ISSN 1367-5435

PDF banerjee2000.pdf - Published Version Restricted to Registered users only Download (88Kb) | Request a copy

Abstract

An extracellular b-galactosidase which catalyzed the production of galacto-oligosaccharide from lactose was harvested from the late stationary-phase of Bacillus sp MTCC 3088. The enzyme was purified 36.2-fold by ZnCl2 precipitation, ion exchange, hydrophobic interaction and gel filtration chromatography with an overall recovery of 12.7%. The molecular mass of the purified enzyme was estimated to be about 484 kDa by gel filtration on a Sephadex G- 200 packed column and the molecular masses of the subunits were estimated to be 115, 86.5, 72.5, 45.7 and 41.2 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The isoelectric point of the native enzyme, determined by polyacrylamide gel electrofocusing, was 6.2. The optimum pH and temperature were 8 and 60°C, respectively. The Michaelis–Menten constants determined with respect to o-NO2-phenyl-b-D-galactopyranoside and lactose were 6.34 and 6.18 mM, respectively. The enzyme activity was strongly inhibited (68%) by galactose, the end product of lactose hydrolysis reaction. The b-galactosidase was specific for b-D anomeric linkages. Enzyme activity was significantly inhibited by metal ions (Hg2+, Cu2+ and Ag+) in the 1–2.5 mM range. Mg2+ was a good activator. Catalytic activity was not affected by the chelating agent EDTA.

Item Type:	Article
Additional Information:	Copyright of this article belongs to Springer Science.
Uncontrolled Keywords:	purification; extracellular b-galactosidase; o-NO2-phenyl-b-D-galactopyranoside; lactose; Bacillus sp
Subjects:	Q Science > QR Microbiology
Depositing User:	Dr. K.P.S.Sengar
Date Deposited:	02 Feb 2012 16:31
Last Modified:	02 Feb 2012 16:31
URI:	http://crdd.osdd.net/open/id/eprint/849

Actions (login required)

View Item