Interaction of the eighth component of guinea pig complement (C8) with the membrane-bound C5b-7 complex. The binding site of C8 to C5b-7 is formed on association of the alpha-gamma subunit with the beta-chain.

The eighth component (c8) of guinea pig complement consists of three polypeptide chains, the alpha-, beta-, and gamma-chains with M.W. of 60,000, 60,000, and 24,000, respectively. The alpha- and gamma-chains are bound by a disulfide bond(s) forming an alpha-gamma subunit, which is linked noncovalently to the beta-chain. The alpha-gamma subunit and the beta-chain were separated and purified from C8 by treatment with sodium dodecyl sulfate (SDS) and gel chromatography on Sephacryl S-300 in the presence of SDS. After removal of SDS, neither alpha-gamma nor beta showed the hemolytic activity of C8 when assayed independently, but showed significant activity in combination, indicating reconstitution of active C8. The recovery of hemolytic activity was 3.48%. When alpha-gamma and beta were incubated successively with EAC-7 with intervening reconstitution of active C8 on the cells was insignificant, irrespective of the order of the reactions. alpha-gamma and beta did not bind to EAC-7 when added separately, but after recombination 7% of alpha-gamma and 9% of beta bound to EAC-7 when EAC-7 was in excess. These results indicate that the binding site of guinea pig C8 to the membrane-bound C5b-7 complex does not exist on either alpha-gamma or beta only but stretches over both or is formed on one subunit after recombination of the subunits.