title: Characterization of the interaction between Robo1 and heparin and other glycosaminoglycans.
abstract: Roundabout 1 (Robo1) is the cognate receptor for secreted axon guidance molecule, Slits, which function to direct cellular migration during neuronal development and angiogenesis. The Slit2-Robo1 signaling is modulated by heparan sulfate, a sulfated linear polysaccharide that is abundantly expressed on the cell surface and in the extracellular matrix. Biochemical studies have further shown that heparan sulfate binds to both Slit2 and Robo1 facilitating the ligand-receptor interaction. The structural requirements for heparan sulfate interaction with Robo1 remain unknown. In this report, surface plasmon resonance (SPR) spectroscopy was used to examine the interaction between Robo1 and heparin and other GAGs and determined that heparin binds to Robo1 with an affinity of ~650 nM. SPR solution competition studies with chemically modified heparins further determined that although all sulfo groups on heparin are important for the Robo1-heparin interaction, the N-sulfo and 6-O-sulfo groups are essential for the Robo1-heparin binding. Examination of differently sized heparin oligosaccharides and different GAGs also demonstrated that Robo1 prefers to bind full-length heparin chains and that GAGs with higher sulfation levels show increased Robo1 binding affinities.
authors.affiliation: Department of Chemical and Biological Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY 12180, USA.
authors.forename: Fuming
authors.initials: F
authors.lastname: Zhang
authors.forename: Heather A
authors.initials: HA
authors.lastname: Moniz
authors.forename: Benjamin
authors.initials: B
authors.lastname: Walcott
authors.forename: Kelley W
authors.initials: KW
authors.lastname: Moremen
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Linhardt
authors.forename: Lianchun
authors.initials: L
authors.lastname: Wang
mesh_terms: D006025:Glycosaminoglycans / Q000737:chemistry* / Q000378:metabolism; D006493:Heparin / Q000737:chemistry / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000378:metabolism; D009844:Oligosaccharides / Q000378:metabolism; D011971:Receptors, Immunologic / Q000737:chemistry* / Q000378:metabolism; D020349:Surface Plasmon Resonance; D000097905:Roundabout Proteins
chemical_list: D006025:Glycosaminoglycans; D006493:Heparin; D009419:Nerve Tissue Proteins; D009844:Oligosaccharides; D011971:Receptors, Immunologic; D000097905:Roundabout Proteins
keywords: Binding; CSA; CSB; CSC; CSD; CSE; Dis-DS; GAG; HA; HS; Heparin; RU; Robo1; SA; SPR; Slit; Surface plasmon resonance; chondroitin sulfate A; chondroitin sulfate B; chondroitin sulfate C; chondroitin sulfate D; chondroitin sulfate E; degree of polymerization; dermatan disulfate; dp; glycosaminoglycan; heparan sulfate; hyaluronic acid; resonance unit; roundabout 1; secreted axon guidance molecules; streptavidin; surface plasmon resonance