RNA-dependent RNA polymerase from heterobasidion RNA virus 6 is an active replicase in vitro

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http://hdl.handle.net/10138/334913

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Levanova , A A , Vainio , E J , Hantula , J & Poranen , M M 2021 , ' RNA-dependent RNA polymerase from heterobasidion RNA virus 6 is an active replicase in vitro ' , Viruses , vol. 13 , no. 9 , 1738 . https://doi.org/10.3390/v13091738

Title: RNA-dependent RNA polymerase from heterobasidion RNA virus 6 is an active replicase in vitro
Author: Levanova, Alesia A.; Vainio, Eeva J.; Hantula, Jarkko; Poranen, Minna M.
Other contributor: University of Helsinki, Molecular and Integrative Biosciences Research Programme
University of Helsinki, Biosciences
University of Helsinki, Molecular and Integrative Biosciences Research Programme


Date: 2021-09
Language: eng
Number of pages: 20
Belongs to series: Viruses
ISSN: 1999-4915
DOI: https://doi.org/10.3390/v13091738
URI: http://hdl.handle.net/10138/334913
Abstract: Heterobasidion RNA virus 6 (HetRV6) is a double-stranded (ds)RNA mycovirus and a member of the recently established genus Orthocurvulavirus within the family Orthocurvulaviridae. The purpose of the study was to determine the biochemical requirements for RNA synthesis catalyzed by HetRV6 RNA-dependent RNA polymerase (RdRp). HetRV6 RdRp was expressed in Escherichia coli and isolated to near homogeneity using liquid chromatography. The enzyme activities were studied in vitro using radiolabeled UTP. The HetRV6 RdRp was able to initiate RNA synthesis in a primer-independent manner using both virus-related and heterologous single-stranded (ss)RNA templates, with a polymerization rate of about 46 nt/min under optimal NTP concentration and temperature. NTPs with 2′-fluoro modifications were also accepted as substrates in the HetRV6 RdRp-catalyzed RNA polymerization reaction. HetRV6 RdRp transcribed viral RNA genome via semi-conservative mechanism. Furthermore, the enzyme demonstrated terminal nucleotidyl transferase (TNTase) activity. Presence of Mn2+ was required for the HetRV6 RdRp catalyzed enzymatic activities. In summary, our study shows that HetRV6 RdRp is an active replicase in vitro that can be potentially used in biotechnological applications, molecular biology, and biomedicine.
Subject: 1182 Biochemistry, cell and molecular biology
DsRNA virus
Mycovirus
Replicase activity
Semi-conservative transcription
TNTase activity
Viral RdRp
DOUBLE-STRANDED-RNA
MINUS-STRAND
PROTEIN
MECHANISM
viral RdRp
dsRNA virus
DE-NOVO INITIATION
BACTERIOPHAGE-PHI-6
mycovirus
semi-conservative transcription
ROTAVIRUS VP1
MESSENGER-RNA
STRUCTURAL BASIS
replicase activity
TRANSFERASE-ACTIVITY
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