Methylamine reaction and denaturation-dependent fragmentation of complement component 3. Comparison with alpha2-macroglobulin

Abstract

Complement protein C-3 can covalently incoporate [14C]methylamine with a stoichiometry of 0.85 +/- 0.11 mol/mol of protein. The reactive site is located in the larger, Mr = 135,000 peptide subunit of C-3. The methylamine is incorporated as a derivative of glutamic acid, viz. as gamma-glutamylmethylamide, which was identified by high performance liquid chromatography and low resolution mass spectroscopy. C-3 was shown to undergo a specific, denaturation-dependent protein fragmentation in sodium dodecyl sulfate at 90 degrees C. The cleavage results in the partial conversion of the Mr = 135,000 subunit to fragments of Mr = 84,000 and 53,000. The cleavage is completely prevented by reaction of C-3 with methylamine prior to the 90 degrees C incubation. The site of methylamine incorporation (the glutamyl residue) and the peptide fragmentation reaction have been reported for alpha2-macroglobulin (Howard, J.B., Vermeulen, M., and Swenson, R. (1980) J. Biol. Chem. 255, 3820-3823). A comparison of the results for the two proteins suggests that they have a common reactive site.