Identification of a nickel(II) binding site on hemoglobin which confers susceptibility to oxidative deamination and intramolecular cross-linking

Abstract

Complexation of Ni(II) with native state recombinant hemoglobin is shown to produce NH2-terminal deamination and globin cross-linking in the presence of the oxidant potassium peroxymonosulfate (Oxone(TM)). Both the oxidative deamination and cross-linking are exclusive to the β chains. Recombinant hemoglobin mutants have been created to identify protein sequence requirements for these reactions. It was found that His-2 of the β globin is required for redox active Ni(II) complexation, oxidative deamination, and cross-linking. The oxidative deamination results in the formation of a free carbonyl in place of the NH2-terminal amine of the β chain. Most cross- linking of the β globin occurs intramolecularly, forming β globin dimers. Structural characterization of the β globin dimers indicates the presence of heterogeneous cross-links within the central hemoglobin cavity between the NH2 terminus of one β chain and the COOH-terminal region of the other.