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Enzyme Research
Volume 2010, Article ID 132148, 7 pages
Research Article

Biochemical Characterization of a Thiol-Activated, Oxidation Stable Keratinase from Bacillus pumilus KS12

Department of Microbiology, University of Delhi, South Campus, New Delhi 110021, India

Received 23 January 2010; Accepted 16 June 2010

Academic Editor: Roberto Fernandez Lafuente

Copyright © 2010 Rinky Rajput et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


An extracellular keratinase from Bacillus pumilus KS12 was purified by DEAE ion exchange chromatography. It was a 45 kDa monomer as determined by SDS PAGE analysis. It was found to be an alkaline, serine protease with pH and temperature optima of 10 and 60 , respectively. It was thiol activated with two- and eight-fold enhancement in presence of 10 mM DTT and -mercaptoethanol, respectively. In addition, its activity was stimulated in the presence of various surfactants, detergents, and oxidizing agents where a nearly 2- to 3-fold enhancement was observed in presence of and . It hydrolyzed broad range of complex substrates including feather keratin, haemoglobin, fibrin, casein,and -keratin. Analysis of amidolytic activity revealed that it efficiently cleaved phenylalanine leucine alanine- p-nitroanilides. It also cleaved insulin B chain between - , - and - residues.