Nonequilibrium Dynamics Simulations of Nitric Oxide Release: Comparative Study of Nitrophorin and Myoglobin

Nitrophorin 4 (NP4) is a heme protein that reversibly binds nitric oxide (NO), with release rates modulated
by pH change. High-resolution structures of NP4 revealed that pH changes and NO binding induce a large
conformational rearrangement in two loops that serve to protect the heme-bound NO molecule from solvent.
We used extended (110 ns) molecular dynamics simulations of NP4 at pH 5 and pH 7, modeled by selective
deprotonation of acidic groups. Conformational and dynamic changes were observed, consistent with those
found in the crystal. Further, major solvent movement and NO escape were observed at pH 7, while the
ligand remained in the heme binding pocket at pH 5. As a control, we also performed molecular dynamics
(MD) simulations of sperm whale myoglobin, where NO migration into the interior cavities of the protein
was observed, consistent with previous reports. We constructed a kinetic model of ligand escape to quantitatively
relate the microscopic rate constants to the observed rates, and tested the predictions against the experimental
data. The results suggest that release rates of diatomic molecules from heme proteins can be varied by several
orders of magnitude through modest adjustments in geminate rebinding and gating behavior.