Carbohydrate structures of the third component of rat complement. Presence of both high-mannose and complex type oligosaccharide chains.

Abstract

We investigated the carbohydrate structure of the third component of complement (C3) newly synthesized by cultured rat hepatocytes. When the cells were incubated with [3H]mannose, [3H]galactose or [3H]glucosamine, these radioactive precursors were incorporated only into the alpha subunit of C3, demonstrating that only the alpha subunit contains oligosaccharide chains. [3H]Mannose-labelled C3 was purified from the culture medium by immunoaffinity chromatography. Oligosaccharides prepared by Pronase digestion and strong alkaline hydrolysis were separated into two fractions by Bio-Gel P-2 chromatography (Fractions I and II). The two fractions were analysed by concanavalin A-Sepharose chromatography, ion-exchange high-performance liquid chromatography, and Bio-Gel P-4 gel filtration before and after sequential exoglycosidase digestions. It was found that Fraction I contained two complex type oligosaccharide chains, (NeuAc)2(Gal)2(GlcNAc)2(Man)3(GlcNAc)2 and (NeuAc)3(Gal)3(GlcNAc)3(Man)3(GlcNAc)2, and Fraction II contained the high-mannose type, consisting mainly of (Man)8(GlcNAc)2. Taken together with the carbohydrate composition of rat serum C3, the results suggest that rat C3 has one high-mannose type oligosaccharide chain and two complex type chains in the alpha subunit, which is different from the proposal for human C3.