title: Mutational analysis of Kaposica reveals that bridging of MG2 and CUB domains of target protein is crucial for the cofactor activity of RCA proteins.
abstract: The complement system has evolved to annul pathogens, but its improper regulation is linked with diseases. Efficient regulation of the system is primarily provided by a family of proteins termed regulators of complement activation (RCA). The knowledge of precise structural determinants of RCA proteins critical for imparting the regulatory activities and the molecular events underlying the regulatory processes, nonetheless, is still limited. Here, we have dissected the structural requirements of RCA proteins that are crucial for one of their two regulatory activities, the cofactor activity (CFA), by using the Kaposi's sarcoma-associated herpesvirus RCA homolog Kaposica as a model protein. We have scanned the entire Kaposica molecule by sequential mutagenesis using swapping and site-directed mutagenesis, which identified residues critical for its interaction with C3b and factor I. Mapping of these residues onto the modeled structure of C3b-Kaposica-factor I complex supported the mutagenesis data. Furthermore, the model suggested that the C3b-interacting residues bridge the CUB (complement C1r-C1s, Uegf, Bmp1) and MG2 (macroglobulin-2) domains of C3b. Thus, it seems that stabilization of the CUB domain with respect to the core of the C3b molecule is central for its CFA. Identification of CFA-critical regions in Kaposica guided experiments in which the equivalent regions of membrane cofactor protein were swapped into decay-accelerating factor. This strategy allowed CFA to be introduced into decay-accelerating factor, suggesting that viral and human regulators use a common mechanism for CFA.
authors.affiliation: Complement Biology Laboratory, National Centre for Cell Science, Pune 411007, India;
authors.forename: Avneesh Kumar
authors.initials: AK
authors.lastname: Gautam
authors.affiliation: Complement Biology Laboratory, National Centre for Cell Science, Pune 411007, India;
authors.forename: Yogesh
authors.initials: Y
authors.lastname: Panse
authors.affiliation: Department of Biotechnology, S. P. Pune University, Pune 411007, India;
authors.forename: Payel
authors.initials: P
authors.lastname: Ghosh
authors.affiliation: Complement Biology Laboratory, National Centre for Cell Science, Pune 411007, India;
authors.forename: Malik Johid
authors.initials: MJ
authors.lastname: Reza
authors.affiliation: Microbial Containment Complex, National Institute of Virology, Pune 411021, India.
authors.forename: Jayati
authors.initials: J
authors.lastname: Mullick
authors.affiliation: Complement Biology Laboratory, National Centre for Cell Science, Pune 411007, India; arvindsahu@nccs.res.in.
authors.forename: Arvind
authors.initials: A
authors.lastname: Sahu
mesh_terms: D003179:Complement C3b / Q000737:chemistry* / Q000235:genetics; D017244:Complement Factor I / Q000737:chemistry* / Q000235:genetics; D019288:Herpesvirus 8, Human / Q000737:chemistry* / Q000235:genetics; D006801:Humans; D008958:Models, Molecular; D046912:Multiprotein Complexes / Q000737:chemistry* / Q000235:genetics; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D014764:Viral Proteins / Q000737:chemistry* / Q000235:genetics
chemical_list: C473771:KSHV complement control protein, Herpesvirus 8; D046912:Multiprotein Complexes; D014764:Viral Proteins; D003179:Complement C3b; C493611:CFI protein, human; D017244:Complement Factor I
keywords: Kaposica; RCA protein; cofactor activity; complement regulation; immune evasion