Probing the structure of C1 with an anti-C1s monoclonal antibody: the possible existence of two forms of C1 in solution.
Anti-human C1s monoclonal antibody H1532, a mouse gamma-1-immunoglobulin elicited by a C1r2C1s2 immunogen, appeared to bind to the beta-domain of C1s by electron microscopy. In agreement with this observation, Western blotting demonstrated good binding to unreduced C1s, but no binding to the alpha or gamma-B domains. When added to solutions of the C1r2C1s2 tetramer, HI532 converted the 8.7 S tetramer into an 18 S complex, which was seen by electron microscopy to be a dimer of parallel C1s x C1r x C1r x C1s molecules cross-linked by two bivalent monoclonal antibodies. If increasing amounts of HI532 were added to C1r2C1s2 followed by addition of equivalent C1q, there was a progressive loss of hemolytic activity, which became zero when two equivalents of antibody HI532 were added. When two equivalents of HI532 were added to serum or C1 reconstituted overnight from purified subcomponents, there was an immediate loss of approximately 50% of the hemolytic activity; thereafter, activity decayed slowly and even after 24 hr, 10-30% of the activity remained. The rapid loss of only 50% of the activity would be readily explained by the existence of two conformations of C1, one of which was rapidly disassembled by antibody, and the other was resistant to disassembly. These two conformations may correspond to two previously proposed structures for the C1 complex.
Tseng
Y
Y
Molecular Biology Institute and the Department of Chemistry and Biochemistry, University of California, Los Angeles 90095, U.S.A.
Phillips
M L
ML
Schumaker
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VN
D000818:Animals; D000911:Antibodies, Monoclonal; D015153:Blotting, Western; D003172:Complement C1 / Q000276:immunology*; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008854:Microscopy, Electron; D012996:Solutions
D000911:Antibodies, Monoclonal; D003172:Complement C1; D012996:Solutions