Effects of site-directed mutagenesis in the N-terminal domain of thermolysin on its stabilization.

ABSTRACT: The thermolysin variant G8C/N60C/S65P in which the triple mutation in the N-terminal domain, Gly8→Cys/Asn60→Cys/Ser65→Pro, is undertaken increases stability [Yasukawa, K. and Inouye, K. (2007) Improving the activity and stability of thermolysin by site-directed mutagenesis. Biochim. Biophys. Acta 1774, 1281-1288] and its mechanism is examined in this study. The apparent denaturing temperatures based on ellipticity at 222 nm of the wild-type thermolysin (WT), G8C/N60C, S65P and G8C/N60C/S65P were 85, >95, 88 and >95°C, respectively. The first-order rate constants, k(obs), of the thermal inactivation of WT and variants at 10 mM CaCl₂ increased with increasing thermal treatment temperatures (70-95°C), and those at 80°C decreased with increasing CaCl₂ concentrations (1-100 mM). The k(obs) valu

SUBMITTER: Kawasaki Y 

PROVIDER: S-EPMC3528001 | biostudies-literature | 2013 Jan

REPOSITORIES: biostudies-literature