The Relationship between Enzyme Conformational Change, Proton Transfer, and Phosphoryl Transfer in β‑Phosphoglucomutase

Molecular details
for the timing and role of proton transfer in
phosphoryl transfer reactions are poorly understood. Here, we have
combined QM models, experimental NMR measurements, and X-ray structures
to establish that the transition of an archetypal phosphoryl transfer
enzyme, βPGM, from a very closed near-attack conformation to
a fully closed transition state analogue (TSA) conformation triggers
both partial proton transfer from the general acid–base residue
to the leaving group oxygen and partial dissociation of the transferring
phosphoryl group from the leaving group oxygen. Proton transfer continues
but is not completed throughout the reaction path of the phosphoryl
transfer with the enzyme in the TSA conformation. Moreover, using
interacting quantum atoms (IQA) and relative energy gradient (REG)
analysis approaches, we observed that the change in the position of
the proton and the corresponding increased electrostatic repulsion
between the proton and the phosphorus atom provide a stimulus for
phosphoryl transfer in tandem with a reduction in the negative charge
density on the leaving group oxygen atom. The agreement between solution-phase 19F NMR measurements and equivalent QM models of βPGMWT and βPGMD10N TSA complexes confirms the
protonation state of G6P in the two variants, validating the employed
QM models. Furthermore, QM model predictions of an AlF4 distortion in response to the proton position are confirmed using
high resolution X-ray crystal structures, not only providing additional
validation to the QM models but also further establishing metal fluorides
as highly sensitive experimental predictors of active-site charge
density distributions.