Each of the three binding sites on complement factor H interacts with a distinct site on C3b.
Factor H (fH) restricts activation of the alternative pathway of complement at the level of C3, both in the fluid phase and on self-structures, but allows the activation to proceed on foreign structures. To study the interactions between fH and C3b we used surface plasmon resonance analysis (Biacore(R)) and eight recombinantly expressed fH constructs containing fragments of the 20 short consensus repeat domains (SCRs) of fH. We analyzed the binding of these constructs to C3b and its cleavage products C3c and C3d. Three binding sites for C3b were found on fH. Site 1 was localized to the five amino-terminal SCRs (SCR1-5), and its reciprocal binding site on C3b was found to be lost upon the cleavage of C3b to C3c and C3d. Site 2 on fH was localized by exclusion probably within or near SCRs 12-14 (fragment SCR8-20 bound to C3b, C3c, and C3d; SCR8-11 did not bind to C3b at all; and SCR15-20 bound only to the C3d part of C3b). Site 3 on fH for C3b was localized to the carboxyl-terminal SCRs 19-20, and its reciprocal binding site was mapped to the C3d part of C3b. In conclusion, we confirmed and mapped three binding sites on fH for C3b and demonstrated that the three binding sites on fH interact with distinct sites on C3b. Multiple reciprocal interactions between C3b and fH can provide a basis for the different reactivity of the alternative pathway with different target structures.
Jokiranta
T S
TS
Department of Bacteriology and Immunology, Haartman Institute and HD-Diagnostics, Haartmaninkatu 3, University of Helsinki, Finland. sakari.jokiranta@helsinki.fi
Hellwage
J
J
Koistinen
V
V
Zipfel
P F
PF
Meri
S
S
D001665:Binding Sites; D015374:Biosensing Techniques; D003001:Cloning, Molecular; D003179:Complement C3b / Q000737:chemistry* / Q000378:metabolism*; D017242:Complement Factor H / Q000737:chemistry* / Q000378:metabolism*; D016384:Consensus Sequence; D006801:Humans; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D020449:Repetitive Sequences, Amino Acid; D020349:Surface Plasmon Resonance / Q000379:methods
C493495:CFH protein, human; D010446:Peptide Fragments; D011994:Recombinant Proteins; D003179:Complement C3b; D017242:Complement Factor H