Activation Heat Capacities in Pyridoxal Phosphate Enzymes

Much attention has been given to the temperature dependence
of
enzyme catalyzed reaction rates in the last ∼100 years. Over
the last couple of decades, it has become apparent that activity decreases
before enzymes lose their structural integrity. Two viable models
have been proposed to account for this behavior. In one, an inactive
conformation(s) becomes more populated at higher temperatures. In
the other, a difference in heat capacity between ground and transition
state conformational ensembles is invoked. Here, the temperature dependence
of the activity of 16 different combinations of pyridoxal phosphate
enzymes and substrates has been measured. All show non-Arrhenius activity
vs temperature profiles. These are generally best accommodated by
the second model where the ΔG‡ term in the Eyring equation is expanded in terms of ΔH‡, ΔS‡, and ΔCp‡ (macromolecular
rate theory (MMRT) equation). The values of ΔCp‡ extracted by curve fitting are negative
and in the range of −1 to −2 kJ/(mol K). Several data
sets fit better to a modified MMRT equation that additionally includes
the temperature dependence of ΔCp‡ (dΔCp‡/dT). The values of dΔCp‡/dT from curve fitting
are also negative. The additional dΔCp‡/dT term has major effects on
temperature profiles at the low and high ends of the biological temperature
range. Molecular dynamics simulations using the AMBER-FB15 force field
were performed on a set of six model proteins and two of the enzymes
studied experimentally. Convex energy vs temperature relationships
are observed, which require a positive dCp/dT term to fit the data well. The simulations for
ground and transition state structures for aspartate aminotransferase
and d-serine deaminase allow calculation of ΔCp‡ and dΔCp‡/dT for comparison
to experiments. Negative values for ΔCp‡ and dΔCp‡/dT are obtained for both enzymes.
The simulations generally reproduce well the experimental values of
ΔCp‡ but the dΔCp‡/dT values
are overestimated. The results support a model in which a large, looser
ensemble of ground state-compatible conformations is in equilibrium
with a very small, tighter transition state-optimized ensemble that
allows bond making and breaking to occur with a low energetic barrier.