Dynamical properties of alpha-amylase in the folded and unfolded state: the role of thermal equilibrium fluctuations for conformational entropy and protein stabilisation

A comparative analysis of thermal equilibrium fluctuations occurring in a m esophilic and in a thermophilic alpha -amylase was performed to study the e ffect of structural fluctuations on thermostability. The thermal fluctuatio ns determining the conformational entropy of both enzymes have been charact erised for the folded (at 30 degreesC and 60 degreesC) and for the unfolded state by applying neutron spectroscopy (at 30 degreesC). The folded state shows a higher structural flexibility for the thermophilic protein as compa red to the mesophilic homologue. In contrast, the unfolded state of both en zymes is rather similar with respect to the structural fluctuations. On the basis of this result, a mechanism characterised by entropic stabilisation (i.e., smaller DeltaS for the unfolding transition of thermophilic a-amylas e) can be assumed to be responsible for the higher thermostability of the t hermophilic enzyme. (C) 2001 Elsevier Science B.V. All rights reserved.