Thermal adaptation of extremozymes: Temperature-sensitive contact analysis of serine proteases.

Abstract

Enzyme thermal adaptation reflects a delicate interplay between the sequence, structure, and dynamics of proteins which are fined tuned to meet environmental demands of organisms. Understanding these evolutionary relationships can drive advances in bioengineering, including the design of industrial enzymes and the development of novel therapeutics. This work explores sequence-to-dynamics connections in subtilisin-like serine protease homologs using a recently developed computational methodology that employs expanded ensemble simulations and temperature-sensitive contact analysis. We reveal that thermophilic enzymes achieve thermal stability through extensive salt bridges and hydrophobic networks, whereas psychrophilic enzymes rely on the stability of targeted interactions for cold adaptation. An unsupervised cluster analysis of residue conservation, flexibility, and hydrophobic interactions provides a comprehensive view of residue-specific contributions to thermal adaptation. These findings highlight the coordinated roles of conserved and variable regions in enzyme stability, offering a framework for tailoring enzymes to specific thermal properties for biotechnological applications.