title: Molecular interactions between MASP-2, C4, and C2 and their activation fragments leading to complement activation via the lectin pathway.
abstract: Activation of component C3 is central to the pathways of complement and leads directly to neutralization of pathogens and stimulation of adaptive immune responses. The convertases that catalyze this reaction assemble from fragments of complement components via multistep reactions. In the lectin pathway, mannose-binding lectin (MBL) and ficolins bind to pathogens and activate MBL-associated serine protease-2 (MASP-2). MASP-2 cleaves C4 releasing C4a and generating C4b, which attaches covalently to the pathogen surface upon exposure of its reactive thioester. C2 binds to C4b and is also cleaved by MASP-2 to form the C3 convertase (C4b2a). To understand how this complex process is coordinated, we have analyzed the interactions between MASP-2, C4, C2, and their activation fragments and have compared MASP-2-catalyzed cleavage of C4b2 and C2. The data show that C2 binds tightly to C4b but not to C4, implying that C4 and C2 do not circulate as preformed complexes but that C2 is recruited only after prior activation of C4. Following cleavage of C4, C4b still binds to MASP-2 (KD approximately 0.6 microM) and dissociates relatively slowly (koff approximately 0.06 s-1) compared with the half-life of the thioester (<or=0.7 s) (Sepp, A., Dodds, A. W., Anderson, M. J., Campbell, R. D., Willis, A. C., and Law, S. K. (1993) Protein Sci. 2, 706-716). We propose that the C4b.MASP-2 interaction favors attachment of C4b near to the activating MBL.MASP complex on the bacterial surface so that, following recruitment of C2, the proximity of enzyme and substrate (C4b2) combined with more favorable reaction kinetics drive the formation of the C3 convertase, promoting complement activation.
authors.affiliation: Department of Infection, Immunity, and Inflammation, University of Leicester, Leicester LE1 9HN, United Kingdom. rw73@le.ac.uk
authors.forename: Russell
authors.initials: R
authors.lastname: Wallis
authors.forename: Alister W
authors.initials: AW
authors.lastname: Dodds
authors.forename: Daniel A
authors.initials: DA
authors.lastname: Mitchell
authors.forename: Robert B
authors.initials: RB
authors.lastname: Sim
authors.forename: Kenneth B M
authors.initials: KB
authors.lastname: Reid
authors.forename: Wilhelm J
authors.initials: WJ
authors.lastname: Schwaeble
mesh_terms: D000818:Animals; D016466:CHO Cells; D002384:Catalysis; D003167:Complement Activation; D003175:Complement C2 / Q000378:metabolism*; D050577:Complement C3-C5 Convertases / Q000378:metabolism; D003181:Complement C4 / Q000378:metabolism*; D006224:Cricetinae; D003412:Cricetulus; D007700:Kinetics; D037102:Lectins / Q000737:chemistry / Q000378:metabolism*; D050606:Mannose-Binding Protein-Associated Serine Proteases / Q000502:physiology*; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry; D020349:Surface Plasmon Resonance
chemical_list: D003175:Complement C2; D003181:Complement C4; D037102:Lectins; D011994:Recombinant Proteins; D050577:Complement C3-C5 Convertases; D050606:Mannose-Binding Protein-Associated Serine Proteases; C491338:Masp2 protein, rat