Purification and characterization of organic solvent-stable protease from organic solvent-tolerant Pseudomonas aeruginosa PST-01

• Published in: Journal of bioscience and bioengineering 1999, Vol.87 (1), p.61-68
• Main Authors: Ogino, Hiroyasu, Watanabe, Fumitake, Yamada, Mitsuharu, Nakagawa, Satoshi, Hirose, Toshiya, Noguchi, Atsushi, Yasuda, Masahiro, Ishikawa, Haruo
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Biology of microorganisms of confirmed or potential industrial interest; Biotechnology; Fundamental and applied biological sciences. Psychology; metalloprotease; Miscellaneous; Mission oriented research; organic solvent-stable enzyme; organic solvent-tolerant microorganism; PROTEASAS; PROTEASE; PROTEASES; PSEUDOMONAS AERUGINOSA; PURIFICACION; PURIFICATION; SOLVANT; SOLVENTES; SOLVENTS
• ISSN: 1389-1723; 1347-4421
• DOI: 10.1016/S1389-1723(99)80009-7

An organic solvent-stable protease (PST-01 protease) in a culture broth of organic solvent-tolerant Pseudomonas aeruginosa PST-01 was purified by successive hydrophobic interaction chromatography using Butyl-Toyopearl gels. The purified enzyme was homogeneous as determined by SDS-polyacrylamide gel electrophoresis. PST-01 protease had a molecular mass of 38 kDa. The optimum temperature and pH for casein hydrolysis were 55°C and 8.5, respectively. PST-01 protease was stable at pH 8–12 and below 50°C and was determined to be a metalloprotease which was inhibited by EDTA, 1,10-phenanthroline, and phosphoramidon. PST-01 protease inhibited by EDTA was reactivated completely by the addition of zinc or cobalt ions. The stability of PST-01 protease in solutions containing water-soluble organic solvents or alcohols was higher than that in the absence of organic solvent. Furthermore, in general, PST-01 protease was more stable than commercially available proteases, namely, subtilisin Carlsberg, thermolysin, and α-chymotrypsin, in the presence of water-soluble organic solvents or alcohols.