TRIMETHYLPHOSPHINE BINDING TO HORSE-HEART AND SPERM-WHALE MYOGLOBINS - KINETICS, PROTON MAGNETIC-RESONANCE ASSIGNMENT AND NUCLEAR OVERHAUSER EFFECT INVESTIGATION OF THE HEME POCKET

Two-dimensional nuclear magnetic resonance techniques have been used t o assign resonances corresponding to the heme pocket and several other residues of horse heart and sperm whale myoglobins ligated by trimeth ylphosphine. The assignment procedure was based mainly on the nuclear Overhauser effect connectivities with the ligand and the heme substitu ents. For quantitative measurements of Overhauser effects, application of truncated driven techniques between a proton from distal residues and methyl groups from the ligand was used to determine internuclear d istances. These new results have permitted us to map the heme pockets and to investigate the conformational differences in the heme pockets between horse heart and sperm whale myoglobins. The interproton distan ces between distal amino acid residues and trimethylphosphine were fou nd to be longer in horse heart myoglobin relative to those in sperm wh ale myoglobin. This result suggests that the size of the heme pocket i s larger in horse heart myoglobin.Association and dissociation rate co nstants were measured for trimethylphosphine binding to myoglobins. Bo th values were four times larger for horse heart myoglobin than those for sperm whale myoglobin. This observation confirms the structural re sults obtained with NMR studies and is rationalized by a greater stabi lization of a larger pocket in horse heart myoglobin relative to sperm whale myoglobin.