Transmembrane channel formation by complement: functional analysis of the number of C5b6, C7, C8, and C9 molecules required for a single channel.
Earlier studies have shown that sequential treatment of resealed erythrocyte ghosts with C5b6, C7, C8, and C9 leads to insertion of hydrophobic peptides from these complement proteins into the membrane and assembly of transmembrane channels. The number of molecules of each of the proteins required for assembly of the membrane-associated channel structure was evaluated by measuring the quantitative relationship between the doses of the individual proteins and the release of two trapped markers, sucrose and inulin, from ghosts after channel formation. The incubation period was sufficient to attain equilibrium of marker distribution between the ghosts and the extracellular fluid. Two markers of different size (sucrose and inulin, 0.9 and 3 nm molecular diameter, respectively) were used in order to develop information on the molecular composition of small and large channels, respectively. We found that participation of C5b6, C7, and C8 in channel formation displayed one-hit characteristics, regardless of marker size. By contrast, the participation of C9 was one-hit with respect to the sucrose marker, whereas with respect to the inulin marker the C9 reaction was multi-hit. Our results are compatible with the view that these markers are released through a channel structure in the membrane that is a monomer of C5b--9 of the composition C5b61 C71C81C9n, in which n = 1 for channels permitting passage of sucrose and n = 2 for channels allowing transit of inulin.
D000818:Animals; D003182:Complement C5 / Q000378:metabolism; D050776:Complement C5b; D003184:Complement C7 / Q000378:metabolism; D003185:Complement C8 / Q000378:metabolism; D003186:Complement C9 / Q000378:metabolism; D003165:Complement System Proteins / Q000378:metabolism*; D004306:Dose-Response Relationship, Immunologic; D004910:Erythrocyte Membrane / Q000276:immunology; D007473:Ion Channels; D046911:Macromolecular Substances; D012756:Sheep
D003182:Complement C5; D003184:Complement C7; D003185:Complement C8; D003186:Complement C9; D007473:Ion Channels; D046911:Macromolecular Substances; D050776:Complement C5b; D003165:Complement System Proteins