Resonance fluorescence spectra of Zn-substituted myoglobin

Laser-induced fluorescence spectra, including narrow resonance lines, were measured for Zn-substituted myoglobin (ZnMb) in the lowest optical absorption band at 4 K by using a combination of short light pulses from a CW mode-locked laser for the excitation and a time-correlated single-photon counting method for the detection. The site-energy distribution function, i.e., the distribution of the number of chromophore at various sites as a function of the zero-phonon transition energy, was determined from the excitation profile of the narrow resonance fluorescence line. The single-site fluorescence spectrum was determined by use of the saturation effect of laser-induced fluorescence. By comparing the laser-induced fluorescence spectra and the absorption spectrum with the spectra calculated by using the experimentally determined site-energy distribution and single-site fluorescence spectra, we found that the coupling between the electrons of the chromophore and the vibrations of the polypeptide was quite weak. Furthermore, the density of states of vibrational modes of myoglobin weighted by the coupling strength between the chromophore and the polypeptide chain was determined from the single-site fluorescence spectrum by solving an integral equation numerically.