Myoglobin-CO Substate Structures and Dynamics:  Multidimensional Vibrational Echoes and Molecular Dynamics Simulations

Abstract

Spectrally resolved infrared stimulated vibrational echo data were obtained for sperm whale carbonmonoxymyoglobin (MbCO) at 300 K. The measured dephasing dynamics of the CO ligand are in agreement with dephasing dynamics calculated with molecular dynamics (MD) simulations for MbCO with the residue histidine-64 (His64) having its imidazole epsilon nitrogen protonated (N(epsilon)-H). The two conformational substate structures B(epsilon) and R(epsilon) observed in the MD simulations are assigned to the spectroscopic A(1) and A(3) conformational substates of MbCO, respectively, based on the agreement between the experimentally measured and calculated dephasing dynamics for these substates. In the A(1) substate, the N(epsilon)-H proton and N(delta) of His64 are approximately equidistant from the CO ligand, while in the A(3) substate, the N(epsilon)-H of His64 is oriented toward the CO, and the N(delta) is on the surface of the protein. The MD simulations show that dynamics of His64 represent the major source of vibrational dephasing of the CO ligand in the A(3) state on both femtosecond and picosecond time scales. Dephasing in the A(1) state is controlled by His64 on femtosecond time scales, and by the rest of the protein and the water solvent on longer time scales.

Ultrafast Dynamics of Myoglobin without the Distal Histidine: Stimulated Vibrational Echo Experiments and Molecular Dynamics Simulations

“…In contrast, the spectrum of wtMbCO shows at least three distinct absorption lines, centered at 1934 (A 3 ), 1945 (A 1 ), and 1965 cm -1 (A 0 ). 5,26,27 The A 0 peak, the smallest of the three features at room temperature and neutral pH, is believed to arise from configurations in which the ligand interacts most weakly with H64 because the distal histidine is swung out of the pocket. [36][37][38][39]56 As has been discussed in detail previously 5,8,[57][58][59] Figure 1 suggests that the substitution of the distal histidine (H64) by a valine spectroscopically mimics the A 0 state of wtMbCO.…”

“…The experimental setup is similar to that described previously. 5 Tunable mid-IR pulses with a center frequency of 1967 cm -1 were generated by an optical parametric amplifier pumped with a regeneratively amplified Ti:Sapphire laser. The nearly transform-limited Gaussian-shaped pulses had a bandwidth and pulse duration of 130 cm -1 and 120 fs, respectively.…”

“…5,8,10,11,52 In this picture, the force exerted by the local electric field on the electric dipole of the CO induces a shift in the CO transition frequency. Therefore, the frequency fluctuates in time with the dynamics of the local electric field.…”

“…The combination of complementary spectroscopic and computational studies can provide a detailed and self-consistent picture of protein structure, dynamics, and function. 3,5,[8][9][10][11] The spectrally resolved infrared stimulated vibrational echo has been successfully applied to investigate protein structure and dynamics. 6,7,9,12,13 Application of this technique to heme proteins has focused on the CO bound to the active site.…”

“…Comparison of measured and simulated absorption spectra and vibrational echoes showed that the primary structural difference between A 1 and A 3 states could be assigned to a rotation of the singly protonated imidazole group of the distal histidine, H64. 5,8,10,11 A wealth of experimental and theoretical evidence underscores the importance of H64 in myoglobin function and dynamics. 23,[30][31][32][33] H64 is the most polar residue near the active site, and interacts strongly with both CO and O 2 ligands.…”