Structural characterization of a zinc high-affinity binding site in rhodopsin.

For the first time to our knowledge, X-ray absorption spectroscopy (XAS) has been used to investigate the environment of putative Zn(2+) binding sites in rhodopsin. We studied native purified nondeionized rhodopsin without any further addition of Zn(2+), as well as with 1.5 mol of Zn(2+)-as zinc chloride-per mole of protein. Three different binding sites in rhodopsin were considered based on computational chemistry studies, and a quantitative analysis of the XAS signal was performed by fitting the experimental data to their simulated XAS spectra. Our results demonstrate that Zn(2+) is intrinsically bound to rhodopsin and are compatible with the existence of an octahedral coordination involving six oxygen atoms in the first shell (average Zn-O distance of 2.08 A), and with a second coordination shell containing one or two phosphorus or sulfur atoms at an average distance of 2.81 A.