PERTURBATIONS OF THE DISTAL HEME POCKET IN HUMAN MYOGLOBIN MUTANTS PROBED BY INFRARED-SPECTROSCOPY OF BOUND CO - CORRELATION WITH LIGAND-BINDING KINETICS

The infrared spectra of CO bound to human myoglobin and myoglobin muta nts at positions His-64, Val-68, Asp-60, and Lys-45 on the distal side have been measured between 100 and 300 K. Large differences are obser ved with mutations at His-64 and Val-68 as well as with temperature an d pH. Although distal His-64 is found to affect CO bonding, Val-68 als o plays a major role. The variations are analyzed qualitatively in ter ms of a simple model involving steric interaction between the bound CO and the distal residues. A strong correlation is found between the fi nal barrier height to CO recombination and the CO stretch frequency: a s compared to wild type, the barrier is smaller in those mutants that have a higher CO stretch frequency (nu(CO)) and vice versa. Possible r easons for this correlation are discussed. It is emphasized that the t emperature and pH dependence of both the kinetics and the infrared spe ctra must be measured to obtain a consistent picture.