Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 1991 Jun;65(6):2993-3000.
doi: 10.1128/JVI.65.6.2993-3000.1991.

Expression, purification, and properties of recombinant encephalomyocarditis virus RNA-dependent RNA polymerase

S Sankar  1 , A G Porter
Affiliations

Expression, purification, and properties of recombinant encephalomyocarditis virus RNA-dependent RNA polymerase

S Sankar et al. J Virol. 1991 Jun.

Abstract

Encephalomyocarditis (EMC) virus RNA-dependent RNA polymerase was expressed in Escherichia coli as a fusion protein with glutathione S-transferase (GST), which allowed easy purification of the fusion protein by affinity chromatography on immobilized glutathione. Inclusion of a thrombin cleavage site between the GST carrier and the viral enzyme facilitated the release of purified mature EMC virus RNA polymerase from the GST carrier by proteolysis with thrombin. The purified recombinant enzyme has a molecular mass of about 52 kDa and is recognized by polyclonal immune serum raised against a peptide sequence corresponding to the C-terminal region of the protein. The recombinant enzyme comigrates with immunoprecipitated EMC virus RNA polymerase from infected mouse L929 cell extracts when run in parallel lanes on a sodium dodecyl sulfate-polyacrylamide gel. The enzyme exhibits rifampin-resistant, poly(A)-dependent poly(U) polymerase activity and RNA polymerase activity, which are both oligo(U) dependent. Template-size products are synthesized in in vitro reactions with EMC virus genomic RNA or globin mRNA. The availability of recombinant EMC virus RNA polymerase in a purified form will allow biochemical analysis of its role in the replication of the virus as well as structure-function studies of this unique class of enzyme.

PubMed Disclaimer

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Eur J Biochem. 1989 Sep 1;184(1):39-45 - PubMed
    1. J Virol. 1986 Oct;60(1):43-53 - PubMed
    1. Gene. 1988 Jul 15;67(1):31-40 - PubMed
    1. Virology. 1988 Jun;164(2):301-8 - PubMed
    1. J Biol Chem. 1985 Jun 25;260(12):7628-35 - PubMed

MeSH terms

LinkOut - more resources