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

Abstract

Recent studies have shown that Ni(II) in human blood serum is bound to albumin and the binding site is located at the NH 2 -terminus of the protein. In this article, the nature of the Ni(II)-binding site of the NH 2 -terminal tripeptide segment of human albumin: L-aspartyl-L-alanyl-L-histidine-N-methyl amide (AAHNMA) is reported as deduced by 13 C- and 1 H-nmr spectroscopy. The 13 C 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 NH 2 group. The introduction of Ni(II) ion into the solution containing peptide causes profound changes in the 1 H-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 13 C spin–lattice relaxation times (T 1 ) 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 NH 2 -terminal peptide of human albumin involving the α-NH 2 , N(3) imidazole, two deprotonated peptide nitrogens, and Asp COO − group in a pentacoordinated structure.