title: Mapping of three major heparin-binding sites on laminin and identification of a novel heparin-binding site on the B1 chain.
abstract: Laminin, a major basement membrane glycoprotein, interacts with many basement membrane- and cell surface-associated heparin-like macromolecules. In order to understand these interactions better, we have tried to map heparin-binding sites on laminin precisely. Electron microscopy revealed three major heparin-binding sites: 1) on the globule of the long arm; 2) on the outer globule of the short arms; and 3) on the inner globule of the short arms. Elution of heparin bound to a laminin affinity column with a linear salt gradient produced three peaks at 0.15, 0.17, and 0.20 M NaCl. When the laminin-heparin interaction was examined in the presence of increasing salt concentrations by the technique of rotary shadowing, the weakest binding was assigned to the inner globule of the short arms and the strongest to the globule of the long arm. One peptide termed AC15, with the sequence Arg-Ile-Gln-Asn-Leu-Leu-Lys-Ile-Thr-Asn-Leu-Arg-Ile-Lys-Phe-Val-Lys from the B1 chain, was identified as a heparin-binding sequence localized on the outer globule of the lateral short arm. Because the two stronger heparin-binding sites were mapped in domains participating in laminin self-association, the effect of heparin on this phenomenon was examined using turbidity and electron microscopy. At low heparin concentrations, laminin oligomer and polymer formation was slightly enhanced. At high heparin concentrations, a drastic inhibition of polymerization was observed, and laminin was detected to be mainly monomeric in rotary-shadowed samples. These results suggest that local variation in the concentration of heparin-like macromolecules might be a crucial factor in determining the association of matrix macromolecules and therefore the structure of basement membranes.
authors: Department of Laboratory Medicine and Pathology, University of Minnesota Medical School, Minneapolis 55455.
authors: K
authors: K
authors: Kouzi-Koliakos
authors: G G
authors: GG
authors: Koliakos
authors: E C
authors: EC
authors: Tsilibary
authors: L T
authors: LT
authors: Furcht
authors: A S
authors: AS
authors: Charonis
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002846:Chromatography, Affinity; D006493:Heparin / Q000302:isolation & purification / Q000378:metabolism*; D007797:Laminin / Q000302:isolation & purification / Q000378:metabolism* / Q000648:ultrastructure; D046911:Macromolecular Substances; D051379:Mice; D008969:Molecular Sequence Data; D009374:Neoplasms, Experimental / Q000378:metabolism
chemical_list: D007797:Laminin; D046911:Macromolecular Substances; D006493:Heparin