OLIGOSACCHARIDE SEQUENCE OF HUMAN BREAST-CANCER CELL HEPARAN-SULFATE WITH HIGH-AFFINITY FOR LAMININ

Laminin-1 is a basement membrane glycoprotein implicated in tumor-host adhesion, which involves the cell-binding domain(s) of laminin-l and tumor cell surface heparan sulfate (HS), The specific tumor cell surfa ce HS oligosaccharide sequences that are necessary for binding to lami nin-l have not been characterized. To identify this laminin-binding ol igosaccharide sequence, Glc-NSO4-rich oligosaccharides terminating wit h [H-3]2,5-anhydromannitol (AMan(R)) residues were isolated from human breast cancer cell (MCF-7)-derived HS through hydrazinolysis/high pH (4.0) nitrous acid treatment/[H-3]NaBH4 reduction. These oligosacchari des were chromatographed on a laminin-l affinity column, A high affini ty dodecasaccharide was isolated and characterized. Disaccharide analy sis yielded IdoA(2-SO4) --> AMan(R)(6-SO4) as the only disaccharide up on treatment of this dodecasaccharide with nitrous acid at low pH (1.5 ), The sequence of laminin-binding high affinity oligosaccharide is th erefore [IdoA(2-SO4 --> GlcNSO(4)(6-SO4)](5)[IdoA(2-SO4) --> AMan(R)(6 -SO4)]. Low affinity dodecasaccharides composed of [IdoA(2-SO4) --> Gl cNSO(4)(6-SO4)](5), [IdoA(2-SO4) --> GlcNSO(4)] were also isolated by laminin-l affinity chromatography, Molecular modeling studies indicate that a heparin-binding peptide sequence corresponding to amino acid r esidues 3010-3031 (KQNCLSSRASFRGCVRNLRLSR) in the G domain of laminin- l, modeled as a right-handed alpha-helix, carries an array of basic re sidues well placed to bind to clusters of sulfate groups on the high a ffinity dodecasaccharide.