title: A carboxy terminal domain of the L protein of rinderpest virus possesses RNA triphosphatase activity - The first enzyme in the viral mRNA capping pathway.
abstract: The large protein L of negative-sense RNA viruses is a multifunctional protein involved in transcription and replication of genomic RNA. It also possesses enzymatic activities involved in capping and methylation of viral mRNAs. The pathway for mRNA capping followed by the L protein of the viruses in the Morbillivirus genus has not been established, although it has been speculated that these viruses may follow the unconventional capping pathway as has been shown for some viruses of Rhabdoviridae family. We had earlier shown that the large protein L of Rinderpest virus expressed as recombinant L-P complex in insect cells as well as the ribonucleoprotein complex from purified virus possesses RNA triphosphatase (RTPase) and guanylyltransferase activities, in addition to RNA dependent RNA polymerase activity. In the present work, we demonstrate that RTPase as well as nucleoside triphosphatase (NTPase) activities are exhibited by a subdomain of the L protein in the C terminal region (a.a. 1640-1840). The RTPase activity depends absolutely on a divalent cation, either magnesium or manganese. Both the RTPase and NTPase activities of the protein show dual metal specificity. Two mutant proteins having alanine mutations in the glutamic acid residues in motif-A of the RTPase domain did not show RTPase activity, while exhibiting reduced NTPase activity suggesting overlapping active sites for the two enzymatic functions. The RTPase and NTPase activities of the L subdomain resemble those of the Vaccinia capping enzyme D1 and the baculovirus LEF4 proteins.
authors.affiliation: Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Piyush Kumar
authors.initials: PK
authors.lastname: Singh
authors.affiliation: Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Nivedita
authors.initials: N
authors.lastname: Ratnam
authors.affiliation: Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Kannan Boosi
authors.initials: KB
authors.lastname: Narayanarao
authors.affiliation: Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Harigopalarao
authors.initials: H
authors.lastname: Bugatha
authors.affiliation: Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Anjali A
authors.initials: AA
authors.lastname: Karande
authors.affiliation: Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India. Electronic address: shaila@mcbl.iisc.ernet.in.
authors.forename: Shaila
authors.initials: S
authors.lastname: Melkote Subbarao
mesh_terms: D017766:Acid Anhydride Hydrolases / Q000737:chemistry / Q000378:metabolism*; D000595:Amino Acid Sequence; D008969:Molecular Sequence Data; D012315:RNA Caps; D012367:RNA, Viral / Q000378:metabolism*; D012302:Rinderpest virus / Q000737:chemistry*; D017386:Sequence Homology, Amino Acid; D014764:Viral Proteins / Q000737:chemistry / Q000378:metabolism*
chemical_list: D012315:RNA Caps; D012367:RNA, Viral; D014764:Viral Proteins; D017766:Acid Anhydride Hydrolases; C023090:RNA triphosphatase
keywords: Metalloenzyme; Negative strand RNA viruses; RNA modification; RNA triphosphatase; Rinderpest virus L protein; mRNA capping