Mutant and wild-type myoglobin-CO protein dynamics: Vibrational echo experiments

Picosecond infrared vibrational echo experiments on a mutant protein, H64V myoglobin-CO, are described and compared to experiments on wild-type myoglobin-CO. H64V is myoglobin with the distal histidine replaced by a valine. The vibrational dephasing experiments examine the influence of protein dynamics on the CO ligand, which is bound to the active site of the mutant protein, from low temperature to physiologically relevant temperatures. The experiments were performed with a mid-infrared free electron laser tuned to the CO stretch mode at 1969 cm(-1). The vibrational echo results are combined with infrared pump-probe measurements of the CO vibrational lifetime to yield the homogeneous pure dephasing. The homogeneous pure dephasing is the Fourier transform of the homogeneous line width with the lifetime contribution removed. The measurements were made from 60 to 300 K and show that the CO vibrational spectrum is inhomogeneously broadened at all temperatures studied. The mutant protein's CO vibrational pure dephasing rate is similar to 20% slower (narrower homogeneous pure dephasing line width) than the wild-type protein at all temperatures, although the only difference between the two proteins is the replacement of the wild-type's polar distal histidine amino acid by a nonpolar valine. These results provide insights into the mechanisms of the transmission of protein fluctuations to the CO ligand bound at the active site, and they are consistent with previously proposed mechanisms of protein-ligand coupling.