title: Bivalent and co-operative binding of complement factor H to heparan sulfate and heparin.
abstract: FH (Factor H) with 20 SCR (short complement regulator) domains is a major serum regulator of complement, and genetic defects in this are associated with inflammatory diseases. Heparan sulfate is a cell-surface glycosaminoglycan composed of sulfated S-domains and unsulfated NA-domains. To elucidate the molecular mechanism of binding of FH to glycosaminoglycans, we performed ultracentrifugation, X-ray scattering and surface plasmon resonance with FH and glycosaminoglycan fragments. Ultracentrifugation showed that FH formed up to 63% of well-defined oligomers with purified heparin fragments (equivalent to S-domains), and indicated a dissociation constant K(d) of approximately 0.5 μM. Unchanged FH structures that are bivalently cross-linked at SCR-7 and SCR-20 with heparin explained the sedimentation coefficients of the FH-heparin oligomers. The X-ray radius of gyration, R(G), of FH in the presence of heparin fragments 18-36 monosaccharide units long increased significantly from 10.4 to 11.7 nm, and the maximum lengths of FH increased from 35 to 40 nm, confirming that large compact oligomers had formed. Surface plasmon resonance of immobilized heparin with full-length FH gave K(d) values of 1-3 μM, and similar but weaker K(d) values of 4-20 μM for the SCR-6/8 and SCR-16/20 fragments, confirming co-operativity between the two binding sites. The use of minimally-sulfated heparan sulfate fragments that correspond largely to NA-domains showed much weaker binding, proving the importance of S-domains for this interaction. This bivalent and co-operative model of FH binding to heparan sulfate provides novel insights on the immune function of FH at host cell surfaces.
authors.affiliation: Department of Structural and Molecular Biology, Darwin Building, University College London, Gower Street, London WC1E 6BT, U.K.
authors.forename: Sanaullah
authors.initials: S
authors.lastname: Khan
authors.forename: Ruodan
authors.initials: R
authors.lastname: Nan
authors.forename: Jayesh
authors.initials: J
authors.lastname: Gor
authors.forename: Barbara
authors.initials: B
authors.lastname: Mulloy
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Perkins
mesh_terms: D001665:Binding Sites / Q000235:genetics; D017242:Complement Factor H / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006493:Heparin / Q000378:metabolism*; D006497:Heparan Sulfate / Q000378:metabolism*; D006801:Humans; D008956:Models, Chemical; D011485:Protein Binding / Q000235:genetics; D017433:Protein Structure, Secondary / Q000235:genetics; D053838:Scattering, Small Angle
chemical_list: D017242:Complement Factor H; D006493:Heparin; D006497:Heparan Sulfate; C493495:CFH protein, human