RESUMEN
A decrease of the hemolytic function of human C3 results from reduction with sodium borohydride (NaBH4) or with dithiothreitol (DTT) followed by alkylation with iodoacetamide. In the presence of 3.5 x 10(-2) M NaBH4, the loss of hemolytic activity (68%) is paralleled by a loss of C3b binding to EAC14oxy2 cells (67%), immune adherence (71%), and hemagglutinability (74%), but by no loss of C5 convertase function. Sulfhydryl analysis reveals that two disulfide bonds, both presumed to be situated in the C3 alpha subunit, are broken: one at 2.2 x 10(-2) M and one at 3.5 x 10(-2) M NaBH4. A contrasting inactivation profile is observed in the presence of 1.0 x 10(-1) M DTT. Loss of hemolytic function (76%) is paralleled by a decrease of C5 convertase function (87%), whereas C3b binding, immune adherence, and hemagglutinability are either unaffected or not significantly affected. Sulfhydryl analysis indicates the cleavage of four disulfide bonds; three bonds within C3 alpha and beta are broken at 5.0 x 10(-4) M DTT, and one additional bond within C3 alpha at 1.0 x 10(-3) M DTT. Antigenicity as determined by immunoelectrophoresis is unaltered at any of the above concentrations. The implication of these findings is discussed.