The long-term objective of this research program is to discover and elucidate the structural determinants of heme protein function. Attainment of this objective will provide a better understanding of how the protein can modulate the intrinsic heme reactivity.
The specific aims of the proposed project include determination of the role of the protein in two aspects of the chemical reactivity of the Fe(III) state of heme proteins: (1) the activation of hydrogen peroxide and subsequent cleavage of the oxygen-oxygen bond of the peroxide; and (2) the binding of small exogenous ligands such as cyanide, fluoride, and azide. The experimental design makes use of heme proteins from two different functional classes, the oxygen-transport proteins and the heme enzymes. Mammalian metmyoglobin and G. dibranchiata hemoglobin will be the representatives of the oxygen-transport proteins and cytochrome c peroxidase will be representative of the heme enzymes. The role of specific amino acids in the reaction of the heme proteins with hydrogen peroxide and in ligand binding reactions will be probed using proteins engineered for this purpose through site-directed mutagenesis. The basic strategy is to be able to manipulate the protein structure in order to convert the oxygen-transport proteins into heme proteins with efficient peroxidase activity and to be able to convert cytochrome c peroxidase into a protein with reversible oxygen binding characteristics.
| Vitello, L B; Erman, J E; Miller, M A et al. (1993) Effect of arginine-48 replacement on the reaction between cytochrome c peroxidase and hydrogen peroxide. Biochemistry 32:9807-18 |
| Erman, J E; Vitello, L B; Miller, M A et al. (1993) Histidine 52 is a critical residue for rapid formation of cytochrome c peroxidase compound I. Biochemistry 32:9798-806 |
| Vitello, L B; Erman, J E; Miller, M A et al. (1992) Effect of Asp-235-->Asn substitution on the absorption spectrum and hydrogen peroxide reactivity of cytochrome c peroxidase. Biochemistry 31:11524-35 |
