Structure, Properties, and Engineering of the Major Zinc Binding Site on Human Albumin

Abstract

Most blood plasma zinc is bound to albumin, but the structure of the binding site has not been determined. Zn K-edge extended x-ray absorption fine structure spectroscopy and modeling studies show that the major Zn(2+) site on albumin is a 5-coordinate site with average Zn-O/N distances of 1.98 A and a weak sixth O/N bond of 2.48 A, consistent with coordination to His(67) and Asn(99) from domain I, His(247) and Asp(249) from domain II (residues conserved in all sequenced mammalian albumins), plus a water ligand. The dynamics of the domain I/II interface, thought to be important to biological function, are affected by Zn(2+) binding, which induces cooperative allosteric effects related to those of the pH-dependent neutral-to-base transition. N99D and N99H mutations enhance Zn(2+) binding but alter protein stability, whereas mutation of His(67) to alanine removes an interdomain H-bond and weakens Zn(2+) binding. Both wild-type and mutant albumins promote the safe management of high micromolar zinc concentrations for cells in cultures.