Hydrophobic Residues Regulate Distal Histidine Coordinations in Human Cgb and Ngb

Neuroglobin (Ngb) and Cytoglobin (Cgb) are new members of the vertebrate globin family. The solved structures of Ngb and Cgb indicated that both of them contain hexa-coordinated heme 3, and one of the endogenous ligands should be eliminated at the first step when an exogenous ligand bind with the heme iron. Since the replacement of ligands is important to express physiological functions of these globins, the position of the dissociated histidine and surrounding residues are decisive to the functions. Although several physiological roles have been discussed, they are still unidentified yet. Here we focused on hydrophobic residues which compose heme pocket and are close to the distal histidine. The residues were substituted to less or more bulky ones, and were investigated with spectroscopic methods. In order to study the heme pocket properties, CO and CN(-) affinities were measured for the ferrous and ferric states, respectively, in both Ngb and Cgb. In addition, effects of the introduced mutations were monitored with resonance Raman spectroscopy.