¹H N.M.R Studies of High-Spin Complexes of Soybean Leghemoglobin. Interactions Between the Distal Histidine and Acetate, Formate and Fluoride Ligands

Abstract

High-resolution ¹H n.m.r . spectra have been measured for the acetate, formate and fluoride complexes of ferric soybean leghaemoglobin a. The chemical shifts of the resolved ow-field haem resonances are very sensitive to pH, reflecting titration of a group, assigned as the distal histidine, with pK_a 7.1, 7.3 and 6.2 for the acetate, formate and fluoride complexes respectively. The magnitude of the pH-induced resonance shifts indicates direct interaction between the distal histidine and the axial ligand . The hyperfine shifted resonances of ferrous deoxyleghaemoglobin, in which no sixth ligand is expected, are insensitive to pH over the range pH 5.5-9.3. Evidence for conformational mobility of the distal histidine is presented.