title: Reconstitution of the RNA-dependent RNA polymerase activity of Antheraea mylitta cypovirus in vitro using separately expressed different functional domains of the enzyme.
abstract: Antheraea mylitta cytoplasmic polyhedrosis virus is a segmented dsRNA virus of the family Reoviridae. Segment 2 (S2)-encoded RNA-dependent RNA polymerase (RdRp) helps the virus to propagate its genome in the host cell of the silkworm, Antheraea mylitta. Cloning, expression, purification and functional analysis of individual domains of RdRp have demonstrated that the purified domains interact in vitro. The central polymerase domain (PD) shows nucleotide binding properties, but neither the N-terminal domain (NTD) nor the C-terminal domain (CTD). Isolated PD does not exhibit RdRp activity but this activity can be reconstituted when all three domains are included in the reaction mixture. Molecular dynamics simulation suggests that the isolated PD has increased internal motions in comparison to when it is associated with the NTD and CTD. The motions of the separated PD may lead to the formation of a less accessible RNA template-binding channel and, thus, impair RdRp activity.
authors.affiliation: Department of Biotechnology, Indian Institute of Technology Kharagpur, Kharagpur-721302, India.
authors.forename: Anirban
authors.initials: A
authors.lastname: Kundu
authors.affiliation: Department of Biotechnology, Indian Institute of Technology Kharagpur, Kharagpur-721302, India.
authors.forename: Amlan
authors.initials: A
authors.lastname: Roychowdhury
authors.affiliation: Department of Biotechnology, Indian Institute of Technology Kharagpur, Kharagpur-721302, India.
authors.forename: Madhuparna
authors.initials: M
authors.lastname: Bose
authors.affiliation: Department of Biotechnology, Indian Institute of Technology Kharagpur, Kharagpur-721302, India.
authors.forename: Amit Kumar
authors.initials: AK
authors.lastname: Das
authors.affiliation: Department of Biotechnology, Indian Institute of Technology Kharagpur, Kharagpur-721302, India.
authors.forename: Ananta K
authors.initials: AK
authors.lastname: Ghosh
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D016679:Genome, Viral / Q000235:genetics; D056004:Molecular Dynamics Simulation; D009036:Moths / Q000821:virology*; D017434:Protein Structure, Tertiary / Q000235:genetics; D012367:RNA, Viral / Q000235:genetics*; D012324:RNA-Dependent RNA Polymerase / Q000235:genetics*; D012087:Reoviridae / Q000235:genetics*; D014779:Virus Replication / Q000235:genetics*
chemical_list: D012367:RNA, Viral; D012324:RNA-Dependent RNA Polymerase