J Biol Chem, Vol. 273, Issue 33, 21111-21114, August 14, 1998

Oligosaccharide Sequence of Human Breast Cancer Cell Heparan Sulfate with High Affinity for Laminin

Narayanan Parthasarathy, Lisa F. Gotow, James D. Bottoms, Timothy E. Kute^¶, William D. Wagner, and Barbara Mulloy

From the ^¶ Department of Pathology and  Section of Comparative Medicine, Wake Forest University School of Medicine, Winston-Salem, North Carolina 27157-1040 and the  National Institute for Biological Standards and Control, Blanche Lane, South Mimms, Potters Bar, Harts, En6 3QG, United Kingdom

Laminin-1 is a basement membrane glycoprotein implicated in tumor-host adhesion, which involves the cell-binding domain(s) of laminin-1 and tumor cell surface heparan sulfate (HS). The specific tumor cell surface HS oligosaccharide sequences that are necessary for binding to laminin-1 have not been characterized. To identify this laminin-binding oligosaccharide sequence, GlcNSO₄-rich oligosaccharides terminating with [³H]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/[³H]NaBH₄ reduction. These oligosaccharides were chromatographed on a laminin-1 affinity column. A high affinity dodecasaccharide was isolated and characterized. Disaccharide analysis yielded IdoA(2-SO₄)  AMan_R(6-SO₄) as the only disaccharide upon treatment of this dodecasaccharide with nitrous acid at low pH (1.5). The sequence of laminin-binding high affinity oligosaccharide is therefore [IdoA(2-SO₄)  GlcNSO₄(6-SO₄)]₅[IdoA(2-SO₄)  AMan_R(6-SO₄)]. Low affinity dodecasaccharides composed of [IdoA(2-SO₄)  GlcNSO₄(6-SO₄)]₅, [IdoA(2-SO₄)  GlcNSO₄] were also isolated by laminin-1 affinity chromatography. Molecular modeling studies indicate that a heparin-binding peptide sequence corresponding to amino acid residues 3010-3031 (KQNCLSSRASFRGCVRNLRLSR) in the G domain of laminin-1, modeled as a right-handed -helix, carries an array of basic residues well placed to bind to clusters of sulfate groups on the high affinity dodecasaccharide.