title: The ninth component of human complement (C9). Functional activity of the b fragment.
abstract: The domain structure of human complement protein C9 was investigated by determining the functional activities of the NH2-terminal (C9a) and COOH-terminal (C9b) fragments obtained by cleavage of C9 with alpha-thrombin. The two fragments were separated by preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis and renatured by dialysis against buffers containing zwitterionic detergents. The C9b fragment produced membranolytic activities in three independent assays. First, it produced single, ion-conducting channels of varying conductances in planar lipid membranes. Most of the channels had an average conductance of 11 picoSiemens and an average lifetime of about 30 s. The channels showed lipid specificity and a 3-fold preference for conducting K+ over Na+. Second, the fragment also caused specific marker release from liposomes which was inhibitable by a C9b-specific monoclonal antibody, and third, it lysed erythrocytes in the absence of a fully assembled C5b-8 complex. The isolated C9a fragment did not produce single channels in planar lipid membranes but was also effective in releasing markers from liposomes and in lysing erythrocytes. Secondary structure predictions indicate the presence of several amphiphilic, "surface-seeking" segments in the primary structure of C9 which are mainly alpha-helices in C9b and beta-sheets in C9a. These results may indicate the presence of surface-binding domains in the NH2-terminal half and channel-forming domains in the COOH-terminal portion of native, monomeric C9.
mesh_terms: D000911:Antibodies, Monoclonal; D003186:Complement C9 / Q000032:analysis* / Q000494:pharmacology; D004553:Electric Conductivity; D004591:Electrophoresis, Polyacrylamide Gel; D005452:Fluoresceins / Q000378:metabolism; D006461:Hemolysis; D006801:Humans; D007473:Ion Channels / Q000187:drug effects; D008081:Liposomes / Q000378:metabolism; D008433:Mathematics; D008563:Membrane Lipids / Q000378:metabolism; D010446:Peptide Fragments / Q000032:analysis* / Q000494:pharmacology; D010713:Phosphatidylcholines; D010743:Phospholipids / Q000378:metabolism; D011487:Protein Conformation; D013329:Structure-Activity Relationship
chemical_list: D000911:Antibodies, Monoclonal; D003186:Complement C9; D005452:Fluoresceins; D007473:Ion Channels; D008081:Liposomes; D008563:Membrane Lipids; D010446:Peptide Fragments; D010713:Phosphatidylcholines; D010743:Phospholipids; C024098:6-carboxyfluorescein; C024990:asolectin