title: Structure based modification of Bluetongue virus helicase protein VP6 to produce a viable VP6-truncated BTV.
abstract: Bluetongue virus core protein VP6 is an ATP hydrolysis dependent RNA helicase. However, despite much study, the precise role of VP6 within the viral capsid and its structure remain unclear. To investigate the requirement of VP6 in BTV replication, we initiated a structural and biological study. Multinuclear nuclear magnetic resonance spectra were assigned on his-tagged full-length VP6 (329 amino acid residues) as well as several truncated VP6 variants. The analysis revealed a large structured domain with two large loop regions that exhibit significant conformational exchange. One of the loops (amino acid position 34-130) could be removed without affecting the overall fold of the protein. Moreover, using a BTV reverse genetics system, it was possible to demonstrate that the VP6-truncated BTV was viable in BHK cells in the absence of any helper VP6 protein, suggesting that a large portion of this loop region is not absolutely required for BTV replication.
authors.affiliation: Microbiology & Immunology, Division of Animal Science, Department of Bioresource Science, Graduate School of Agricultural Science, Kobe University, 1-1, Rokkodai, Nada-ku, Kobe-City 657-8501, Japan; Faculty of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine, Keppel Street, London WC1E 7HT, UK.
authors.forename: Eiko
authors.initials: E
authors.lastname: Matsuo
authors.affiliation: Division of Molecular Biosciences, Centre for Structural Biology, Imperial College London, South Kensington, London SW7 2AZ, UK.
authors.forename: Esther
authors.initials: E
authors.lastname: Leon
authors.affiliation: Division of Molecular Biosciences, Centre for Structural Biology, Imperial College London, South Kensington, London SW7 2AZ, UK.
authors.forename: Steve J
authors.initials: SJ
authors.lastname: Matthews
authors.affiliation: Faculty of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine, Keppel Street, London WC1E 7HT, UK. Electronic address: polly.roy@lshtm.ac.uk.
authors.forename: Polly
authors.initials: P
authors.lastname: Roy
mesh_terms: D000818:Animals; D001820:Bluetongue virus / Q000235:genetics* / Q000502:physiology; D006224:Cricetinae; D019906:Nuclear Magnetic Resonance, Biomolecular; D010446:Peptide Fragments / Q000302:isolation & purification / Q000378:metabolism; D020365:RNA Helicases / Q000737:chemistry*; D014758:Viral Core Proteins / Q000737:chemistry*; D014779:Virus Replication
chemical_list: D010446:Peptide Fragments; D014758:Viral Core Proteins; D020365:RNA Helicases
keywords: Bluetongue virus; Multinuclear nuclear magnetic resonance; Reverse genetics; VP6