Protein dynamics. Vibrational coupling, spectral broadening mechanisms, and anharmonicity effects in carbonmonoxy heme proteins studied by the temperature dependence of the Soret band lineshape

Abstract

In this work we study the temperature dependence of the Soret band lineshape of the carbonmonoxy derivatives of sperm whale myoglobin, human hemoglobin, and its isolated alpha and beta subunits. To fit the observed spectral profile we use an analytic expression derived for a system whereby a single electronic transition is coupled to Franck-Condon active vibrational modes, within the adiabatic and harmonic approximation. The vibronic structure of the spectra arises from the coupling with high frequency modes; these modes contribute to the total line shape through a series of Lorentzians with peak positions at vibrational overtones and half width related to the time constant of the population decay of the excited electronic state (homogeneous broadening); moreover, the coupling with low frequency modes broadens each Lorentzian to a Voigtian. Inhomogeneous broadening is modeled as a gaussian distribution of the 0-0 transition frequencies and is therefore added as a constant term to the previous gaussian width. This spectral deconvolution enables us to investigate the different contributions to line broadening and the parameters that characterize the vibrational coupling, as well as their dependence upon protein and solvent composition. The investigation is carried out as a function of temperature in the range 20-300 K; relevant information is obtained by comparing experimental results with theoretical predictions. This work supports a description of the investigated proteins as heterogeneous systems, whose heterogeneity depends on the particular protein and on the composition of the external matrix. The delocalized pi electron cloud of the porphyrin ring is coupled not only to the high frequency vibrational modes of the active site but also to a "bath" of lower frequency modes that involve the entire protein; moreover at suitable temperatures (approximately 200 K), anharmonic motions, which are an obvious prerequisite for the jumping among different conformational substates, become evident.

Structural factors controlling ligand binding to myoglobin: A kinetic hole-burning study

“…The vibrational modes that couple to the electronic transition were taken from resonance Raman data (22,23), as described in ref. 16 for MbCO and in ref. 18 for Mb*CO.…”

“…The spectra of both MbCO and Mb*CO were analyzed for each temperature by using an approach described earlier (16)(17)(18). The method is only outlined here briefly.…”

“…The fitting procedure was described in ref. 16. The vibrational modes that couple to the electronic transition were taken from resonance Raman data (22,23), as described in ref.…”

“…It is a strong absorption band and hence easy to measure, and the underlying electronic transition is well understood (15). The fine structure of the Soret absorption band can be analyzed by taking into account the homogeneous width, the vibronic coupling, and the spectral heterogeneity; important structural information contained in the spectrum can therefore be extracted (16,17). Analysis of the Soret spectra of Mb and Mb*CO and comparison to the known structural data made it possible to quantitatively assign spectral parameters to structural properties (18).…”

Protein dynamics An overview on flash‐photolysis over broad temperature ranges

“…Also a study on the temperature dependence of the Soret light absorption band of sperm whale Mb, adult human Mb, and its CI and p chains demonstrates that the coupling of the porphyrin R+R* electronic transition with highfrequency modes of the tetrapyrrole, detected by resonance Raman spectroscopy, and with low-frequency modes that involve the entire protein, is characterised by strong anharmonic contributions above ca. 180 K [5]. As clearly stated by the authors of this investigation, "anharmonicity of nuclear motions is an obvious prerequisite for jumping among conformational substates of the protein".…”