Electron spin resonance study of the copper(II) complexes of human and dog serum albumins and some peptide analogs

Abstract

Electron spin resonance spectra of the first Cu(II) complexes of human serum albumin, dog serum albumin, l-aspartyl- l-histidine N-methylamide and glycyl-glycyl- l-histidine N-methylamide have been studied using isotopically pure 65Cu in its chloride form. At 77° K, the esr spectra of Cu(II) complex of human serum albumin exhibited only one form of esr signal between pH 6.5 and 11. No intermediate forms were detected. The presence of an equally spaced nine-line superhyperfine structure with spacing ∼15 G indicated considerable covalent bonding between Cu(II) and four nitrogen atoms derived from the protein. The esr spectrum form of Cu(II) bound to human serum albumin detected at neutral pH would be consistent with the participation of four nitrogens from the α-NH 2 group, two peptide groups, and the imidazole group of a histidine residue. In contrast, the esr spectra of Cu(II)-dog serum albumin complex showed a transition from a low pH form to a high pH form as the pH was increased to 9.5. These spectral changes were found to be reversible upon lowering the pH. Ligand superhyperfine splittings in the low pH form of the esr signal of Cu(II)-dog albumin were not resolved. The distinct pH dependence of the esr signals observed in human and dog serum albumin complexes could be correlated to their respective optical spectra changes as a function of pH. At room temperature and in the pH range between 6 and 11, the esr spectra of Cu(II) complexes of l-aspartyl- l-alanyl- l-histidine N-methylamide and glycyl-glycyl- l-histidine N-methylamide exhibited a well-resolved nine-line superhyperfine structure indicating metal coordination with four equivalent nitrogen atoms of peptide.