Effect of exchange amino acid residues of the surface region of the PST-01 protease on its organic solvent-stability

Department of Chemical Engineering, Osaka Prefecture University, 1-1 Gakuen-cho, Naka-ku, Sakai, Osaka 599-8531, Japan.
Biochemical and Biophysical Research Communications (Impact Factor: 2.3). 08/2007; 358(4):1028-33. DOI: 10.1016/j.bbrc.2007.05.047
Source: PubMed


The PST-01 protease from an organic solvent tolerant Pseudomonas aeruginosa has high stability and activity in the presence of various organic solvents. The structure gene of the PST-01 protease was amplified by the error-prone PCR method. The mutated proteases were incubated in the presence of acetonitrile. By measuring remaining activities, two kinds of mutated PST-01 proteases of which the stabilities were changed were selected. These mutations hardly changed the profile of the activity and stability at various pHs. Their activity and stability at higher temperatures were slightly lower than those of the wild-type PST-01 protease. The stabilities of the mutated enzymes in the presence of various organic solvents were greatly reduced. In both the mutated PST-01 proteases, amino acids located at the surface of the enzyme had been substituted.

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    • "Published studies of the mechanism of adaptation of enzymes to function in organic solvent are relatively few. Ogino et al. [116,117] investigated the mechanism of organic solvent tolerance in a Pseudomonas aeruginosa PST-01 protease by site-directed and random mutagenesis. They reported that the disulfide bonds and amino acid residues located on the surface of the molecule play important roles in organic solvent stability of the enzymes. "
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    • "Ghorbel et al. isolated a protease from Bacillus cereus BG1 which retained 89.5 of its original activity, after 15-min incubation at 55°C, in the presence of 2 mM Ca2+; meanwhile, no activity was detected in the absence of Ca2+ [12]. An organic solvent-stable protease from Pseudomonas aeruginosa PST-01 was reported to be stable at the temperature below 50°C [23]. A solvent stable protease, from Pseudomonas aeruginosa PseA retained 80% of its initial activity after heating, for 30 min at 55°C [11]. "
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