title: Analysis of self-association of West Nile virus capsid protein and the crucial role played by Trp 69 in homodimerization.
abstract: The understanding of capsid (C) protein interactions with itself would provide important data on how the core is organized in flaviviruses during assembly. In this study, West Nile (WN) virus C protein was shown to form homodimers using yeast two-hybrid analysis in conjunction with mammalian two-hybrid and in vivo co-immunoprecipitation assays. To delineate the region on the C protein which mediates C-C dimerization, truncation studies were carried out. The results obtained clearly showed that the internal hydrophobic segment flanked by helix I and helix III of WN virus C protein is essential for the self-association of C protein. The crucial role played by Trp 69 in stabilizing the self-association of C protein was also demonstrated by mutating Trp to Gly/Arg/Phe. Substitution of the Trp residue with Gly/Arg abolished the dimerization, whereas substitution with Phe decreased the self-association significantly. The results of this study pinpoint a critical residue in the C protein that potentially plays a role in stabilizing the homotypic interaction.
authors.affiliation: Flavivirology Laboratory, Department of Microbiology, 5 Science Drive 2, National University of Singapore, Singapore 117597, Singapore.
authors.forename: Raghavan
authors.initials: R
authors.lastname: Bhuvanakantham
authors.forename: Mah-Lee
authors.initials: ML
authors.lastname: Ng
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002213:Capsid / Q000737:chemistry*; D036022:Capsid Proteins / Q000737:chemistry*; D002522:Chlorocebus aethiops; D019281:Dimerization; D017353:Gene Deletion; D047468:Immunoprecipitation; D007763:Lac Operon; D016296:Mutagenesis; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D014364:Tryptophan / Q000737:chemistry*; D020798:Two-Hybrid System Techniques; D014709:Vero Cells; D014764:Viral Proteins / Q000737:chemistry; D014902:West Nile virus / Q000378:metabolism*; D001616:beta-Galactosidase / Q000378:metabolism
chemical_list: D036022:Capsid Proteins; D014764:Viral Proteins; D014364:Tryptophan; D001616:beta-Galactosidase