Conformational sampling and kinetics changes across a non-Arrhenius break point in the enzyme thermolysin.

Numerous studies have suggested a significant role that protein dynamics play in optimizing enzyme catalysis, and changes in conformational sampling offer a window to explore this role. Thermolysin from , which is a heat-stable zinc metalloproteinase, serves here as a model system to study changes of protein function and conformational sampling across a temperature range of 16-36 °C. The temperature dependence of kinetics of thermolysin showed a biphasic transition at 26 °C that points to potential conformational and dynamic differences across this temperature. The non-Arrhenius behavior observed resembled results from previous studies of a thermophilic alcohol dehydrogenase enzyme, which also indicated a biphasic transition at ambient temperatures. To explore the non-Arrhenius behavior of thermolysin, room temperature crystallography was applied to characterize structural changes in a temperature range across the biphasic transition temperature. The alternate conformation of side chain fitting to electron density of a group of residues showed a higher variability in the temperature range from 26 to 29 °C, which indicated a change in conformational sampling that correlated with the non-Arrhenius break point.