3D structure of the C3bB complex provides insights into the activation and regulation of the complement alternative pathway convertase

Abstract

Generation of the alternative pathway C3-convertase, the central amplification enzyme of the complement cascade, initiates by the binding of factor B (fB) to C3b to form the proconvertase, C3bB. C3bB is subsequently cleaved by factor D (fD) at a single site in fB, producing Ba and Bb fragments. Ba dissociates from the complex, while Bb remains bound to C3b, forming the active alternative pathway convertase, C3bBb. Using single-particle electron microscopy we have determined the 3-dimensional structures of the C3bB and the C3bBb complexes at Ϸ27Å resolution. The C3bB structure shows that fB undergoes a dramatic conformational change upon binding to C3b. However, the C3b-bound fB structure was easily interpreted after independently fitting the atomic structures of the isolated Bb and Ba fragments. Interestingly, the divalent cationbinding site in the von Willebrand type A domain in Bb faces the C345C domain of C3b, whereas the serine-protease domain of Bb points outwards. The structure also shows that the Ba fragment interacts with C3b separately from Bb at the level of the ␣ NT and CUB domains. Within this conformation, the long and flexible linker between Bb and Ba is likely exposed and accessible for cleavage by fD to form the active convertase, C3bBb. The architecture of the C3bB and C3bBb complexes reveals that C3b could promote cleavage and activation of fB by actively displacing the Ba domain from the von Willebrand type A domain in free fB. These structures provide a structural basis to understand fundamental aspects of the activation and regulation of the alternative pathway C3-convertase.C3 convertase ͉ electron microscopy ͉ factor B

Assembly of macromolecular complexes by satisfaction of spatial restraints from electron microscopy images

“…We further illustrate our method by using experimental negative-stain and cryo-EM class averages as restraints to build accurate models for three additional protein assemblies in the Electron Microscopy Data Bank (EMDB): the human protoconvertase C3bB (EMDB ID code 1583) (19), the bovine mitochondrial supercomplex I 1 III 2 IV 1 (EMDB ID code 1876) (20), and the type I restriction modification complex EcoR124I from E. coli (EMDB ID code 1890) (21) ( Table 1 and Fig. S4).…”

“…The results demonstrate that accurate models for various types of macromolecular assemblies can be obtained by using only a limited number of class averages in combination with other restraints not containing direct 3D information. Next, as an example of the utility of the approach, we use it to compute models for the human transferrin receptor-transferrin (TfR-Tf) complex (18), the human C3bB protoconvertase complex (19), the bovine mitochondrial supercomplex I 1 III 2 IV 1 (20), and the type I restriction modification complex EcoR124I from Escherichia coli (21). Finally, the applicability of the method and possible improvements are discussed.…”

The role of the alternative pathway of complement activation in glomerular diseases

“…Three positively charged amino acids in the MG1 domain (Arg80, Arg72, and Lys82) form the thioester bond with four negative amino acids in the TED domain (Asp1007, Glu1008, Glu1010, Glu1013). The C3F variant, containing neutral glycine (instead of positively charged arginine), has a weaker ability to bind to CFH, which may impair the downregulation of the AP convertase, thus altering the C3 function [14,46,47]. The frequency of F allele varies among different populations, and it is the highest in Caucasian (18%) and the lowest in Asian (1%) [6,48].…”

“…As rs1047286 is located in the MG3 domain of C3 (binding site of CFB), polymorphic variant HAV4-1+ has stronger ability to bind to CFB (Fig. 1) [46,50].…”

“…Carries of C3F allele are at risk for the development of age-related macular degeneration (AMD) [46], partial lipodystrophy (PD) [51], and various types of GN: membranoproliferative glomerulonephritis type II (MPGN II, presently known as dense deposit disease, DDD) [51,52], IgA nephropathy (IgAN) [53,54] and systemic vasculitis (SV) [55,56]. Surprisingly, C3F allele has been shown to have a protective effect on transplant kidney, and its presence in donor's kidney may prolong the graft survival [57].…”