C8 Alpha Research Articles

Effect of reduction on the structural, biologic, and immunologic properties of the third component of human complement.

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.

Characterization of the oligosaccharide units of the fourth component of complement (Ss protein) synthesized by murine macrophages.

The glycosylation of murine C4 (Ss protein) synthesized by peritoneal macrophages has been investigated. Both the intracellular precursor, P-C4 (185), and the C4 alpha- and beta-chains that were secreted into medium were found to be glycosylated; however, no carbohydrate units were detected on the gamma-chain. Analyses of the oligosaccharide units showed that P-C4 (185) appears to contain both a "complex" and a "high mannose" carbohydrate group, the alpha-chain a "complex" group and the beta-chain a "high mannose" carbohydrate unit.