Nickel(II)-binding to the NH2-terminal peptide segment of human serum albumin: 13C-and 1H-nuclear magnetic resonance investigation

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Abstract
Recent studies have shown that Ni(II) in human blood serum is bound to albumin and the binding site is located at the NH2-terminus of the protein. In this article, the nature of the Ni(II)-binding site of the NH2-terminal tripeptide segment of human albumin: L-aspartyl-L-alanyl-L-histidine-N-methyl amide (AAHNMA) is reported as deduced by 13C- and 1H-nmr spectroscopy. The 13C spectrum shows that the Ni(II) complex is in slow exchange on the nmr time scale, and resonances for Ni(II)–peptide complex and metal-free peptide are observed clearly in the pH range 6.4 to 9.1. The Asp COO− carbon is most affected by Ni(II) binding. The imidazole ring system also shows large variation. Furthermore, the results implicate the involvement of the NH2 group. The introduction of Ni(II) ion into the solution containing peptide causes profound changes in the 1H-nmr spectra in the pH range 6.4 to 9.1. Most significantly, there is a complete disappearance of Ala and His N—H protons which confirm the coordination of these two peptide nitrogens. The measurement of the 13C spin–lattice relaxation times (T1) for AAHNMA and its 1/1 complex provide confirmatory evidence for the existence of a definite complex. The overall results suggest that Ni(II) forms complex with the NH2-terminal peptide of human albumin involving the α-NH2, N(3) imidazole, two deprotonated peptide nitrogens, and Asp COO− group in a pentacoordinated structure.