Determination of the binding site of chloro(triethylphosphine)gold(I) in serum albumin

Abstract

Abstract EPR spectroscopy was applied to determine the binding site of chloro(triethylphosphine)gold(I), (ClAuPEt3), in bovine serum albumin (BSA). The EPR spectrum of the spin marker covalently bonded to the sulphydryl group of polypeptide chain shows the presence of two 3-maleimidoPROXYL populations which differ in their way of immobilization. The first population shows the spin marker with a strongly restricted rotation due to hydrogen bonds between the oxygen of the label agent nitroxyl group and hydrogen atoms of the polypeptide chain with possible participation of water. The second population contains the spin marker bound with Cys-34 sulphydryl group located outside the crevice between helices h2 and h3, weakly immobilized by water hydrogen bonds. The addition of ClAuPEt3 results in the changes of the shape of spin labeled albumin EPR spectra. The displacement of 3-maleimidoPROXYL is observed below ClAuPEt3:BSA molar ratio 0.5:1. It is possible that above this molar ratio another bonding of the displaced spin marker takes place, having a high freedom of movement. EPR spectra, recorded after dialysis, show displacement of 3-maleimidoPROXYL molecules and the competition between AuPEt3 and the spin marker to the mutual binding site with cysteine residue.