Volume 6, Number 2, p.p. 51-55
Kinetic and thermodynamic manifestations of the thermally-induced molten-globule-like state of α-chymotrypsin
Michael Shushanyan, Rusudan Sujashvili, Ekaterina Tabuashvili, Maya Makharadze, George Getashvili† and Dimitri E. Khoshtariya
Laboratory of Biochemical Kinetics and Thermodynamics, Institute of Molecular Biology and Biological Physics, Georgian Academy of Sciences, 12 Gotua St, 0160 Tbilisi, Georgia
Calorimetric and kinetic data that indicate the appearance of a temperature-induced molten-globule-like (TIMG) state (in equilibrium) at moderately high temperatures for a model catalytically active protein, α-chymotrypsin, are discussed. The data are rationalized by taking into account the exceptional rôle of solvating (bound) water in shaping the dynamic properties of proteins and their fluctuational flexibility. By heating α-chymotrypsin in its native-like conformation to temperatures approximately 10–15 K below the thermal melting transition (i.e. to 40–45 °C), it is demonstrated that the protein attains a conformation that is still compact and almost native-like but with highly increased fluctuational mobility of the tertiary structure. This is characteristic of a family of molten-globule-like (MGL) states. In microcalorimetric experiments this is manifested through an increased partial heat capacity comparable to that of the urea-induced MGL state, and in kinetic experiments, through the increased apparent Michaelis constant comparable to that observed for the urea-induced MGL state. It is suggested that the TIMG state appears as a result of temperature-induced redistribution of initially strongly bound hydration water, and that the TIMG state is a common form for all globular proteins provided that prior cumulative effects are excluded.
Keywords: α-chymotrypsin, apparent Michaelis constant, enzymatic catalysis, fluctuational mobility, globular proteins, microcalorimetry, partial heat capacity, solvating water, temperature-induced molten-globule-like state, tertiary structure