Analysis of the recognition mechanism of the alternative pathway of complement by monoclonal anti-factor H antibodies: evidence for multiple interactions between H and surface bound C3b.
The ability of the alternative pathway of complement to discriminate targets as either activators or non-activators is mediated by different binding properties of factor H to surface-associated C3b molecules. In the present study we have probed the interaction between H and C3b using five anti-H mAb. The binding sites of the mAb were mapped by Western blotting using both recombinant and trypsin-generated H fragments. Two mAb bound to CCP1 (90X, 196X), two to CCP5 (MRC OX24, 86X) and one to CCP8-15a (131X). At a molar ratio 2:1 of 125I-H:mAb all tested mAb enhanced binding of H to both activator- and non-activator-bound C3b. At higher concentrations two mAb had an inhibitory effect on H binding to surface-associated C3b (OX24, 131X). Thus the mAb 131X inhibits H binding to surface-bound C3b but unlike OX24 it does not bind to the previously described C3b binding site within or near CCP4-5. These results indicate that there is an additional interaction site on factor H for surface-bound C3b.
Jokiranta
T S
TS
Department of Bacteriology and Immunology, Haartman Institute, University of Helsinki, Finland. sjokiran@helsinki.fi
Zipfel
P F
PF
Hakulinen
J
J
Kühn
S
S
Pangburn
M K
MK
Tamerius
J D
JD
Meri
S
S
D000818:Animals; D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D001666:Binding Sites, Antibody; D015153:Blotting, Western; D003179:Complement C3b / Q000032:analysis / Q000276:immunology / Q000378:metabolism*; D017242:Complement Factor H / Q000032:analysis / Q000276:immunology / Q000378:metabolism*; D003170:Complement Pathway, Alternative; D003432:Cross-Linking Reagents; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D010449:Peptide Mapping; D011863:Radioimmunoassay; D011994:Recombinant Proteins / Q000032:analysis / Q000276:immunology / Q000378:metabolism; D014357:Trypsin
D000911:Antibodies, Monoclonal; C493495:CFH protein, human; D003432:Cross-Linking Reagents; D011994:Recombinant Proteins; D003179:Complement C3b; D017242:Complement Factor H; D014357:Trypsin