Purification and characterization of an extreme halothermophilic protease from a halophilic bacterium Chromohalobacter sp. TVSP101

• Autor(a) principal: Vidyasagar,Malashetty
• Data de Publicação: 2009
• Outros Autores: Prakash,S., Mahajan,Vineet, Shouche,Yogesh S., Sreeramulu,K.
• Tipo de documento: Artigo
• Idioma: eng
• Título da fonte: Brazilian Journal of Microbiology
• Texto Completo: http://old.scielo.br/scielo.php?script=sci_arttext&pid=S1517-83822009000100002
• topic: Chromohalobacter sp. TVSP101; halothermophilic protease; purification; organic solvents; osmolytes

An extreme halophilic bacterium was isolated from solar saltern samples and identified based on biochemical tests and 16S r RNA sequencing as Chromohalobacter sp. strain TVSP101. The halophilic protease was purified using ultrafiltration, ethanol precipitation, hydrophobic interaction column chromatography and gel permeation chromatography to 180 fold with 22% yield. The molecular mass of the protease determined by SDS PAGE was 66 kDa. The purified enzyme was salt dependent for its activity and stability with an optimum of 4.5 M NaCl. The optimum temperature for maximum protease activity was 75°C. The protease was optimally active at pH 8 and retained more than 80% of its activity in the range of pH 7-10. Sucrose and glycine at 10% (w/v) were the most effective osmolytes, retained 100% activity in the absence of NaCl. The activity was completely inhibited by ZnCl2 (2 mM), 0.1% SDS and PMSF (1mM). The enzyme was not inhibited by 1mM of pepstatin, EDTA and PCMB. The protease was active and retained 100% it activity in 10% (v/v) DMSO, DMF, ethanol and acetone.