Complement C3 Convertases

Abstract: We have examined the role of macrophage (M phi plasma membrane receptors for the cleaved third complement component (iC3b; CR3) and mannosyl, fucosyl terminated glycoproteins (MFR) in uptake of unopsonized zymosan. Monoclonal antibodies against CR3, M1/70 (Mac-1) and MO1, each inhibited approximately 50% of uptake of 125I-zymosan by murine and human M phi, respectively. Yeast mannan inhibited 0-50% of zymosan uptake in various M phi, in parallel with their expression of MFR. We demonstrated that M phi were the source of C3 in our assay and that the activity of other components of the complement system, namely a C3 convertase, factor I, and a factor I cofactor were also present in serum-free cultures of human monocytes. Macrophage C3 was deposited rapidly, within 10 min, on the zymosan particles and mediated binding, ingestion, and stimulation of superoxide release in BCG-activated and thioglycollate-elicited peritoneal M phi via CR3. Local secretion of complement proteins by M phi themselves can therefore opsonize pathogens and cells able to activate the alternative pathway and effect their destruction.

Abstract: Normal host cells are protected from the destructive action of complement by cell surface complement regulatory proteins In humans, decay-accelerating factor (DAF) and membrane cofactor protein (MCP) play such a biologic role by inhibiting C3 and C5 convertases DAF and MCP accomplish this task by specific mechanisms designated decay-accelerating activity and factor I cofactor activity, respectively In other species, including mice, structural and/or functional homologues of these proteins are not yet well characterized Previous studies have shown that the mouse protein Crry/p65 has certain characteristics of self-protecting complement regulatory proteins For example, Crry/p65 is expressed on a wide variety of murine cells, and when expressed on human K562 erythroleukemic cells, it prevents deposition of mouse C3 fragments on the cell surface during activation of either the classical or alternative complement pathway We have now studied factor I cofactor and decay-accelerating activities of Crry/p65 Recombinant Crry/p65 demonstrates cofactor activity for factor I-mediated cleavage of both mouse C3b and C4b Surprisingly, Crry/p65 also exhibits decay-accelerating activity for the classical pathway C3 convertase strongly and for the alternative pathway C3 convertase weakly Therefore, mouse Crry/p65 uses the specific mechanisms of both human MCP and DAF Although Crry/p65, like MCP and DAF, contains tandem short consensus repeats (SCR) characteristic of C3/C4 binding proteins, Crry/p65 is not considered to be a genetic homologue of either MCP or DAF Thus, Crry/p65 is an example of evolutionary conservation of two specific activities in a single unique protein in one species that are dispersed to individual proteins in another We propose that the repeating SCR motif in this family has allowed this unusual process of evolution to occur, perhaps driven by the use of MCP and DAF as receptors by human pathogens such as the measles virus

Abstract: Two complex enzymes were assembled that both converted C3 to C3b, one consisting of activated properdin (P), native C3, proactivator (PA) and proactivator convertase (PAase), and the other of nephritic factor (NF) and the same three cofactors. By maintaining a critical concentration of PAase, the P-C3 convertase and the NF-C3 convertase were shown to function efficiently without formation of the C3b-feedback enzyme. The former two enzymes are distinct from the C3b-dependent C3 convertase in that they utilize native C3 instead of C3b and PA in an apparently uncleaved form. The P- and NF-C3 convertase express maximal activity within approximately 10 min at 37 degrees C and decay with a half-life of 35 min at 37 degrees C, which is in contradistinction to the reported lability of the C3b-feedback enzyme. P- and NF-C3 convertases are inhibited by their product C3b, which may constitute a heretofore unknown control of the alternative pathway. A direct physical interaction of P with native C3 and C3b was demonstrated by agglutination of C3b-bearing erythrocytes and by agglutination inhibition. Bound C3b thus constitutes the only known receptor of P and may fulfill an important localizing function for P and the P-C3 convertase in vivo. Although P and NF form functionally similar enzymes, they act independently of each other and are apparently immunochemically unrelated proteins.

Abstract: Summary: Evolution of the two gene families of the complement system involved in the formation of the C3 convertases, B/C2 andC3/C4/C5, was studied at the cDNA level in lower vertebrates. Cyclostomes, the most primitive extant vertebrates, seem to possess only one member each of these families, indicating chat gene duplication between B and C2 or among C3, C4 and C5 occurred in the lineage of jawed vertebrates. Typical C3 and C4 cDNAs were identified in both amphibian (Xenopus) and teleost (medaka fish), locating the C3 /C4 gene duplication before the divergence of ray funned fish and lobe-finned Fish, On the other hand, typical B cDNA was identified in Xenopus, whereas teleost counterparts from three species all showed intermediate character between B and C2, suggesting the possibility that the B/C2 gene duplication occurred in die tetrapod lineage. Genetic linkage between these two family genes within the MHC was observed in Xenopus but not in medaka fish.