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J. Biol. Chem., Vol. 275, Issue 23, 17281-17287, June 9, 2000
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From the Program in Cellular Biotechnology, Institute of
Biotechnology, Biocenter Viikki, University of Helsinki,
FIN-00014, Helsinki, Finland
Both genomic and subgenomic RNAs of the
Alphavirus have m7G(5')ppp(5')N (cap0
structure) at their 5' end. Previously it has been shown that
Alphavirus-specific nonstructural protein Nsp1 has
guanine-7N-methyltransferase and guanylyltransferase
activities needed in the synthesis of the cap structure. During normal
cap synthesis the 5'
Identification of a Novel Function of the Alphavirus
Capping Apparatus
RNA 5'-TRIPHOSPHATASE ACTIVITY OF Nsp2*
-phosphate of the nascent viral RNA chain is removed by a specific RNA 5'-triphosphatase before condensation with
GMP, delivered by the guanylyltransferase. Using a novel RNA
triphosphatase assay, we show here that nonstructural protein Nsp2 (799 amino acids) of Semliki Forest virus specifically cleaves the
,
-triphosphate bond at the 5' end of RNA. The same activity was
demonstrated for Nsp2 of Sindbis virus, as well as for the amino-terminal fragment of Semliki Forest virus Nsp2-N (residues 1-470). The carboxyl-terminal part of Semliki Forest virus Nsp2-C (residues 471-799) had no RNA triphosphatase activity. Replacement of
Lys-192 by Asn in the nucleotide-binding site completely abolished RNA
triphosphatase and nucleoside triphosphatase activities of Semliki
Forest virus Nsp2 and Nsp2-N. Here we provide biochemical characterization of the newly found function of Nsp2 and discuss the
unique properties of the entire Alphavirus-capping apparatus.
*
This work was supported by Academy of Finland Grant 8397, Technology Development Center (TEKES), and Center for International Mobility.The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
A Biocentrum Helsinki Fellow. To whom correspondence should be
addressed: Program in Cellular Biotechnology, Institute of Biotechnology, Biocenter Viikki, P. O. Box 56, University of Helsinki, FIN-00014, Helsinki Finland. Tel.: 358-9-191-59400; Fax:
358-9-191-59560; E-mail: leevi.kaariainen@helsinki.fi.
Copyright © 2000 by The American Society for Biochemistry and Molecular Biology, Inc.
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