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Access to the Complement Factor B Scissile Bond Is Facilitated by Association of Factor B with C3b Protein

2011; Elsevier BV; Volume: 286; Issue: 41 Linguagem: Inglês

10.1074/jbc.m111.263418

1083-351X

Dennis E. Hourcade, Lynne M. Mitchell,

Cellular transport and secretion

Factor B is a zymogen that carries the catalytic site of the complement alternative pathway C3 convertase. During convertase assembly, factor B associates with C3b and Mg2+ forming a pro-convertase C3bB(Mg2+) that is cleaved at a single factor B site by factor D. In free factor B, a pair of salt bridges binds the Arg234 side chain to Glu446 and to Glu207, forming a double latch structure that sequesters the scissile bond (between Arg234 and Lys235) and minimizes its unproductive cleavage. It is unknown how the double latch is released in the pro-convertase. Here, we introduce single amino acid substitutions into factor B that preclude one or both of the Arg234 salt bridges, and we examine their impact on several different pro-convertase complexes. Our results indicate that loss of the Arg234-Glu446 salt bridge partially stabilizes C3bB(Mg2+). Loss of the Arg234-Glu207 salt bridge has lesser effects. We propose that when factor B first associates with C3b, it bears two intact Arg234 salt bridges. The complex rapidly dissociates unless the Arg234-Glu446 salt bridge is released whereupon conformational changes occur that activate the metal ion-dependent adhesion site and partially stabilize the complex. The remaining salt bridge is then released, exposing the scissile bond and permitting factor D cleavage. Factor B is a zymogen that carries the catalytic site of the complement alternative pathway C3 convertase. During convertase assembly, factor B associates with C3b and Mg2+ forming a pro-convertase C3bB(Mg2+) that is cleaved at a single factor B site by factor D. In free factor B, a pair of salt bridges binds the Arg234 side chain to Glu446 and to Glu207, forming a double latch structure that sequesters the scissile bond (between Arg234 and Lys235) and minimizes its unproductive cleavage. It is unknown how the double latch is released in the pro-convertase. Here, we introduce single amino acid substitutions into factor B that preclude one or both of the Arg234 salt bridges, and we examine their impact on several different pro-convertase complexes. Our results indicate that loss of the Arg234-Glu446 salt bridge partially stabilizes C3bB(Mg2+). Loss of the Arg234-Glu207 salt bridge has lesser effects. We propose that when factor B first associates with C3b, it bears two intact Arg234 salt bridges. The complex rapidly dissociates unless the Arg234-Glu446 salt bridge is released whereupon conformational changes occur that activate the metal ion-dependent adhesion site and partially stabilize the complex. The remaining salt bridge is then released, exposing the scissile bond and permitting factor D cleavage.