Complement C3b/C3d and cell surface polyanions are recognized by overlapping binding sites on the most carboxyl-terminal domain of complement factor H.
Factor H (FH) is a potent suppressor of the alternative pathway of C in plasma and when bound to sialic acid- or glycosaminoglycan-rich surfaces. Of the three interaction sites on FH for C3b, one interacts with the C3d part of C3b. In this study, we generated recombinant constructs of FH and FH-related proteins (FHR) to define the sites required for binding to C3d. In FH, the C3d-binding site was localized by surface plasmon resonance analysis to the most C-terminal short consensus repeat domain (SCR) 20. To identify amino acids of FH involved in binding to C3d and heparin, we compared the sequences of FH and FHRs and constructed a homology-based molecular model of SCR19-20 of FH. Subsequently, we created an SCR15-20 mutant with substitutions in five amino acids that were predicted to be involved in the binding interactions. These mutations reduced binding of the SCR15-20 construct to both C3b/C3d and heparin. Binding of the wild-type SCR15-20, but not the residual binding of the mutated SCR15-20, to C3d was inhibited by heparin. This indicates that the heparin- and C3d-binding sites are overlapping. Our results suggest that a region in the most C-terminal domain of FH is involved in target recognition by binding to C3b and surface polyanions. Mutations in this region, as recently reported in patients with familial hemolytic uremic syndrome, may lead to indiscriminatory C attack against self cells.
Hellwage
Jens
J
Department of Bacteriology and Immunology, Haartman Institute and Helsinki University Central Hospital, University of Helsinki, Helsinki, Finland. hellwage@pmail.hki-jena.de
Jokiranta
T Sakari
TS
Friese
Manuel A
MA
Wolk
Tobias U
TU
Kampen
Eva
E
Zipfel
Peter F
PF
Meri
Seppo
S
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics / Q000276:immunology; D001053:Apolipoproteins / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites / Q000235:genetics / Q000276:immunology; D001798:Blood Proteins / Q000235:genetics / Q000378:metabolism; D002462:Cell Membrane / Q000276:immunology / Q000378:metabolism; D003179:Complement C3b / Q000378:metabolism*; D015933:Complement C3d / Q000378:metabolism*; D017242:Complement Factor H / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D017124:Conserved Sequence / Q000235:genetics / Q000276:immunology; D006493:Heparin / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D000071228:Polyelectrolytes; D011108:Polymers / Q000378:metabolism*; D011485:Protein Binding / Q000235:genetics / Q000276:immunology; D017434:Protein Structure, Tertiary / Q000235:genetics; D020449:Repetitive Sequences, Amino Acid; D020539:Sequence Analysis, Protein
D001053:Apolipoproteins; D001798:Blood Proteins; D003179:Complement C3b; D015933:Complement C3d; D017242:Complement Factor H; D006493:Heparin; D000071228:Polyelectrolytes; D011108:Polymers; C493495:CFH protein, human; C105970:CFHR3 protein, human; C105092:CFHR4 protein, human; C100631:factor H-related protein 1