Characterisation of the conformational and quaternary structure-dependent heparin-binding region of bovine seminal plasma protein PDC-109

PDC-109, the major heparin-binding protein of bull seminal plasma, binds to sperm choline lipids at ejaculation and modulates capacitation mediated by heparin, Affinity chromatography on heparin-Sepharose showed that polydisperse, but not monomeric, PDC-109 displayed heparin-binding capability. We sought to characterise the surface topology of the quaternary structure-dependent heparin-binding region of PDC-109 by comparing the arginine- and lysine-selective chemical modification patterns of the free and the heparin-bound protein. A combination of reversed-phase peptide mapping of endoproteinase Lys-C-digested PDC-109 derivatives and mass spectrometry was employed to identify modified and heparin-protected residues. PDC-109 contains two tandemly arranged fibronectin type II domains (a, Cys(24)-Cys(61), b, Cys(69)-Cys(109)). The results show that six basic residues (Lys(34), Arg(57), Lys(59), Arg(64), Lys(68), and Arg(104)) mere shielded from reaction with acetic anhydride and 1,2-cyclohexanedione in heparin-bound PDC-109 oligomers, In the H-1-NMR solution structures of single fibronectin type II domains, residues topologically equivalent to PDC-109 Arg(57) (Arg(104)) and Lys(59) lay around beta-strand D on the same face of the domain. In full-length PDC-109, Arg(64) and Lys(68) are both located in the intervening polypeptide between domains a and b, Our data suggest possible quaternary structure arrangements of PDC-109 molecules to form a heparin-binding oligomer, (C) 1999 Federation of European Biochemical Societies.