title: Structural requirements for heparin/heparan sulfate binding to type V collagen.
abstract: Collagen-proteoglycan interactions participate in the regulation of matrix assembly and in cell-matrix interactions. We reported previously that a fragment (Ile824-Pro950) of the collagen alpha1(V) chain, HepV, binds to heparin via a cluster of three major basic residues, Arg912, Arg918, and Arg921, and two additional residues, Lys905 and Arg909 (Delacoux, F., Fichard, A., Cogne, S., Garrone, R., and Ruggiero, F. (2000) J. Biol. Chem. 275, 29377-29382). Here, we further characterized the binding of HepV and collagen V to heparin and heparan sulfate by surface plasmon resonance assays. HepV bound to heparin and heparan sulfate with a similar affinity (KD approximately 18 and 36 nM, respectively) in a cation-dependent manner, and 2-O-sulfation of heparin was shown to be crucial for the binding. An octasaccharide of heparin and a decasaccharide of heparan sulfate were required for HepV binding. Studies with HepV mutants showed that the same basic residues were involved in the binding to heparin, to heparan sulfate, and to the cell surface. The contribution of Lys905 and Arg909 was found to be significant. The triple-helical peptide GPC(GPP)5G904-R918(GPP)5GPC-NH2 and native collagen V molecules formed much more stable complexes with heparin than HepV, and collagen V bound to heparin/heparan sulfate with a higher affinity (in the nanomolar range) than HepV. Heat and chemical denaturation strongly decreased the binding, indicating that the triple helix plays a major role in stabilizing the interaction with heparin. Collagen V and HepV may play different roles in cell-matrix interactions and in matrix assembly or remodeling mediated by their specific interactions with heparan sulfate.
authors.affiliation: Institut de Biologie et Chimie des Protéines, UMR CNRS 5086, Université Claude Bernard Lyon 1, Institut Féderatif de Recherche 128 BioSciences Gerland, 7 Passage du Vercors, 69367 Lyon Cedex 07, Lyon, France.
authors.forename: Sylvie
authors.initials: S
authors.lastname: Ricard-Blum
authors.forename: Mickael
authors.initials: M
authors.lastname: Beraud
authors.forename: Nicolas
authors.initials: N
authors.lastname: Raynal
authors.forename: Richard W
authors.initials: RW
authors.lastname: Farndale
authors.forename: Florence
authors.initials: F
authors.lastname: Ruggiero
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D002417:Cattle; D003094:Collagen / Q000737:chemistry; D024062:Collagen Type V / Q000737:chemistry*; D006224:Cricetinae; D006493:Heparin / Q000737:chemistry*; D006497:Heparan Sulfate / Q000737:chemistry*; D006358:Hot Temperature; D007700:Kinetics; D008969:Molecular Sequence Data; D011485:Protein Binding; D011487:Protein Conformation
chemical_list: D003094:Collagen; D024062:Collagen Type V; D006493:Heparin; D006497:Heparan Sulfate