X-ray absorption spectroscopy proves the trigonal-planar sulfur-only coordination of copper(I) with high-affinity tripodal pseudopeptides

摘要

A series of tripodal ligands L derived from nitrilotriacetic acid (NTA) and extended by three converging metal-binding cysteine chains were previously found to bind selectively copper(I) both in vitro and in vivo. The ligands L~1 (ester) and L~2 (amide) were demonstrated to form copper(I) species with very high affinities, close to that reported for the metal-sequestering metallothioneins (MTs; log K~(Cu-MT) ≈ 19). Here, an in-depth study by Cu K-edge X-ray absorption spectroscopy (XAS) was performed to completely characterize the copper(I) coordination sphere in the complexes, previously evidenced by other physicochemical analyses. The X-ray absorption near-edge structure (XANES) spectra shed light on the equilibrium between a mononuclear complex and a cluster for both L~1 (ester) and L ~2 (amide). The exclusive symmetric CuS_3 geometry adopted in the mononuclear complexes (Cu-S ≈ 2.23 ?) was clearly demonstrated by extended X-ray absorption fine structure (EXAFS) analyses. The EXAFS analyses also proved that the clusters are organized on a symmetric CuS_3 core (Cu-S ≈ 2.26 ?) and interact with three nearby copper atoms (Cu - -Cu ≈ 2.7 ?), consistent with the Cu6S9-type clusters previously characterized by pulsed gradient spin echo NMR spectroscopy. XAS data obtained for other architectures based on the NTA template (L~3 acid, L~4 without a functionalized carbonyl group, etc.) demonstrated the formation of polymetallic species only, which evidence the necessity of the proximal ester or amide group to stabilize the CuS_3 mononuclear species. Finally, XAS was demonstrated to be a powerful method to quantify the equilibrium between the two copper(I) environments evidenced with L~1 and L~2 at different copper concentrations and to determine the equilibrium constants between these two complexes.