The role of the C9b domain in the binding of C9 molecules to EAC1-8 defined by monoclonal antibodies to C9.
Three mAb to human C9, X195, X197, and P40 were used to analyze the roles of the C9a and C9b domains in the reaction of the C9 molecule with sensitized sheep E bearing C1 to C8 (EAC1-8). X195 bound to NH2-terminal (C9a) fragments, and X197 bound to COOH-terminal (C9b) fragments obtained by cleavage of C9 with alpha-thrombin or trypsin. P40 recognized the epitope on the C9b fragment obtained by alpha-thrombin cleavage but did not react with the NH2-terminal or COOH-terminal fragment obtained by trypsin cleavage. In this respect, P40 differed from mAb to C9 reported previously. P40 almost completely inhibited the hemolytic activity of C9. X195 and X197 also inhibited C9 activity, but less effectively than P40. C9 molecules bound to P40 could not bind to EAC1-8 cells. C9 bound to X197 could not bind rapidly to EAC1-8, but prolonged incubation of the C9-X197 complex with EAC1-8 caused considerable lysis of the cells. C9 molecules bound to X195 could bind rapidly to EAC1-8, but their lytic activity was partially inhibited by the bound antibody. From these results, it is concluded that the C9b but not C9a domain contributes to the binding of C9 to EAC1-8 and that the epitope recognized by P40 or a closely adjacent site may be the binding site of C9 molecule to EAC1-8.
Yoden
A
A
Department of Clinical Pathology, Osaka Medical College, Japan.
Moriyama
T
T
Inoue
K
K
Inai
S
S
D000818:Animals; D000911:Antibodies, Monoclonal / Q000502:physiology*; D000937:Antigen-Antibody Reactions; D001666:Binding Sites, Antibody; D001667:Binding, Competitive; D003186:Complement C9 / Q000276:immunology / Q000378:metabolism* / Q000502:physiology*; D015938:Complement Membrane Attack Complex; D003165:Complement System Proteins / Q000378:metabolism*; D005260:Female; D006461:Hemolysis; D006801:Humans; D051379:Mice; D011951:Receptors, Complement / Q000032:analysis*
D000911:Antibodies, Monoclonal; D003186:Complement C9; D015938:Complement Membrane Attack Complex; D011951:Receptors, Complement; C048666:complement C9b; D003165:Complement System Proteins