Excited-State Axial-Ligand Photodissociation and Nonpolar Protein-Matrix Reorganization in Zn(II)-Substituted Cytochromec

Abstract

We have obtained picosecond time-resolved fluorescence spectra from Zn(II)-substituted cytochrome c (ZnCytc) over the 100-ps to 12-ns time scale. The vibronic structure of the Q-band fluorescence spectrum exhibits a pronounced biexponential response (τ1 = 120 ps, τ2 = 7 ns), with the 0−0 band decreasing and the 0−1 band increasing in intensity in a mirror-image fashion with respect to time. These changes evidence a sequential photodissociation of the Zn(II) ion's two axial ligands, which are likely to be supplied by the side chains of the Met 80 and His 18 residues, as in the Fe(II,III)-containing molecule. The time constant for the fast phase of response is significantly slowed when the external solvent contains 50% (v/v) glycerol. These results show that the breaking of the first axial-ligand bond is rate limited by the reorganization of the surrounding protein matrix, which in turn is damped by the surrounding solvent. The protein-matrix response to the axial-ligand photodissociation reaction in ZnCytc is an example of nonpolar solvation dynamics, a reorganization of the protein structure in response to a change in size of an imbedded structure.