Fluid-phase interaction of C1̄ inhibitor (C1̄ Inh) and the subcomponents C1̄ and C1̄s of the first component of complement, C1̄

Abstract

Interactions between proenzymic or activated complement subcomponents of C1̄ and C1̄ Inh (C1̄ inhibitor) were analysed by sucrose-density-gradient ultracentrifugation and sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. The interaction of C1̄ Inh with dimeric C1̄r in the presence of EDTA resulted into two bimolecular complexes accounting for a disruption of C1̄r. The interaction of C1̄ Inh with the Ca2+-dependent C1-r2-C1-s2 complex (8.8 S) led to an 8.5 S inhibited C1̄r-C1̄s-C1̄ Inh complex (1:1:2), indicating a disruption of C1r2 and of C1s2 on C1 Inh binding. The 8.5 S inhibited complex was stable in the presence of EDTA; it was also formed from a mixture of C1̄r, C1̄s and C1̄ Inh in the presence of EDTA or from bimolecular complexes of C1̄r-C1̄ Inh and C1̄s-C1̄ Inh. C1̄r II, a modified C1̄r molecule, deprived of a Ca2+-binding site after autoproteolysis, did not lead to an inhibited tetrameric complex on incubation with C1̄s and C1̄ Inh. These findings suggest that, when C1̄ Inh binds to C1̄r2-C1̄s2 complex, the intermonomer links inside C1̄r2 or C1̄s2 were weakened, whereas the non-covalent Ca2+-independent interaction between C1̄r2 and C1̄s2 is strengthened. The nature of the proteinase-C1̄ Inh link was investigated. Hydroxylamine (1 M) was able to dissociate the complexes partially (pH 7.5) or totally (pH 9.0) when the incubation was performed in denaturing conditions. An ester link between a serine residue at the active site of C1̄r or C1̄s and C1̄ Inh is postulated.