Protein Flexibility and Stability: Thermophiles Know Best

Abstract

Understanding the relation between protein flexibility, stability and function remains one of the most challenging, open questions in biophysical chemistry. For example, proteins need to be flexible to facilitate substrate binding but locally rigid to sustain substrate specificity. Exemplary cases are thermophilic enzymes from archaea and bacteria. These proteins are stable and functional at elevated temperatures but generally lack activity at ambient conditions. Therefore, their thermal stability has been correlated to enhanced mechanical rigidity through the “corresponding states” paradigm [1]. There are, however, a number of studies on thermophilic proteins that have questioned this view [2].In this work we present a comprehensive computational study, that questions the “rigidity paradigm”, at least in its universal character. We compare a pair of homologous G-domain proteins, with their melting temperatures differing by 40 K. Our study points to a clear result: at ambient condition the hyperthermophilic protein has comparable or even enhanced flexibility with respect to the less stable mesophile. When focusing on different time- and length- scales specific behaviors arise. At an atomistic scale, it is found that in the hyperthermophile a more regular alternation of rigid and flexible regions stabilizes a key part of the protein where the unfolding of the mesophile begins. We furthermore find that the conformational landscape of the hyperthermophile is characterized by a higher number of substates, or otherwise an enhanced conformational flexibility that is suggested to broaden its stability curve and raise the melting temperature. We finally compare, for the two proteins, the unfolding paths upon increasing temperature, the kinetic barrier along the early steps of unfolding and the temperature dependency of the stability.[1] A.Wrba, A. Schweiger, V. Schultes, R. Jaenicke, P. Zavodszky, Biochemistry, 1990, 29, 7584-7592.[2] J. Fitter, J. Heberle, Biophys. J, 2000, 79, 1629-1636.