LAUSR

Understanding how enzyme catalysis varies with temperature is key to understanding catalysis itself and, ultimately, how to tune temperature optima. Temperature dependence studies inform on the change in heat capacity during the reaction, ΔCP‡, and we have recently demonstrated that this can expose links between the protein free energy landscape and enzyme turnover. By quantifying ΔCP‡, we capture information on the changes to the distribution of vibrational frequencies during enzyme turnover. The primary experimental tool to probe the role of vibrational modes in a chemical/biological process is isotope effect measurements, since isotopic substitution primarily affects the frequency of vibrational modes at/local to the position of isotopic substitution. We have monitored the temperature dependence of a range of isotope effects on the turnover of a hyper-thermophilic glucose dehydrogenase. We find a progressive effect on the magnitude of ΔCP‡ with increasing isotopic substitution of d-glucose. Our experim...