Improve Salinivibrio zinc-metalloprotease function in less polar organic solvents by increasing surface hydrophobicity.

Organic solvents tend to strip water from protein and thereby disrupt non-covalent forces and decrease enzyme activity and stability. In the present study, we have replaced the surface charge residues in Salinivibrio zinc-metalloprotease (SVP) with hydrophobic ones (E12V, D22I, D24A and D310I) in order to study the effects of surface hydrophobicity with hydrophobic strength of organic solvents. Compared to SVP, D24A exhibited an increase in k and catalytic efficiency and a reduction in thermal inactivation rate in aqueous solvent. Structural studies indicated that the replacement of surface charge residues with hydrophobic residues would not induce conformational changes. C value (the value of solvent concentration where 50% of enzyme activity remains), k (irreversible thermoinactivation rate), and kinetic parameters of E12V, D22I, and D24A were higher in isopropanol and n-propanol. D24A is found to be the most efficient mutant for its remarkable decrease in k value in the presence of isopropanol and n-propanol and a reduction in k value in the presence of dimethylformamide (DMF) and methanol. C value in this variant was increased about 1.2% in DMF, 2% in methanol and isopropanol and 2.5% in n-propanol. Results revealed that, there was a correlation between surface hydrophobicity of SVP and hydrophobic strength of organic solvents.