Post-translational modification of the fourth component of complement. Effect of tunicamycin and amino acid analogs on the formation of the internal thiol ester and disulfide bonds.
The appearance of a functional thiol ester within murine pro-C4 (the intracellular precursor of C4) has been studied. This was assessed by testing the ability of pro-C4 molecules to undergo denaturation-dependent autolytic cleavage. In pulse-chase experiments, [35S]methionine-labeled pro-C4 does not autolyze until approximately 20 min after synthesis by peritoneal macrophages. When intact (not autolyzed) pro-C4 was examined by nonreducing gel electrophoresis, an increase in its apparent Mr was seen, with a time course similar to that for autolysis. Both the capacity to undergo autolytic cleavage and the Mr increase were inhibited by cell culture in the presence of the antibiotic tunicamycin or the threonine analog beta-hydroxynorvaline, both of which inhibit glycosylation. Upon isolation from tunicamycin- or hydroxynorvaline-treated cells, pro-C4 associates with other cell constituents, probably via disulfide bonds. This phenomenon is not seen with the mature (high Mr) form of pro-C4 in control cultures, and can be prevented if the cells are lysed in the presence of a sulfhydryl reagent such as iodoacetamide. These data suggest that the post-translational modification of pro-C4 includes the acquisition of a disulfide-stabilized conformation with a greater apparent Mr. This conformation, along with an intact thiol ester, is necessary for autolytic cleavage to occur.
D000596:Amino Acids / Q000494:pharmacology*; D000818:Animals; D003181:Complement C4 / Q000096:biosynthesis*; D004220:Disulfides / Q000378:metabolism; D005944:Glucosamine / Q000031:analogs & derivatives*; D007700:Kinetics; D008264:Macrophages / Q000378:metabolism; D051379:Mice; D011499:Protein Processing, Post-Translational; D013438:Sulfhydryl Compounds / Q000378:metabolism; D014415:Tunicamycin / Q000494:pharmacology*
D000596:Amino Acids; D003181:Complement C4; D004220:Disulfides; D013438:Sulfhydryl Compounds; D014415:Tunicamycin; D005944:Glucosamine