Dihydrofolate Reductase and the Physical Basis of Enzyme Catalysis
The hallmarks of
catalysis by enzymes are selectivity, specificity, and
speed. However, despite their central role, the physical basis of the
enormous catalytic power of enzymes is not well understood. Initially,
tunnelling was treated through the introduction of a tunnelling
correction to transition-state theory.
However, the examination of the temperature dependence of the kinetic
isotope effects (KIEs) of several enzymatic hydrogen transfer reactions
has
led to a collapse of the semi-classical model for hydrogen tunnelling
and new models were developed to explain these observations such as
environmentally coupled tunnelling in which protein motions are
proposed to drive hydrogen tunnelling.
It is central to our understanding of enzyme catalysis to test these
models further and contrast them with potential alternatives. This is
especially important for the case of temperature dependent KIEs, which
are
consistent with a model where an active promoting motion leads to a
compression of the tunnelling barrier in the reactive state and
enhanced
tunnelling. Alternative models such as multiple conformational states
of
the enzymes also must be examined experimentally.
We use the enzyme dihydrofolate reductase (DHFR) as a model system in
which to study hydride tunnelling. The dependence on temperature, pH,
pressure and solvent of the KIEs from bacterial DHFRs with a range of
optimal temperatures are studied. In addition, DHFR structure and
dynamics are probed using techniques such as circular dichroism and NMR.