Flanegan, James B. and Baltimore, David (1979) Poliovirus Polyuridylic Acid Polymerase and RNA Replicase Have the Same Viral Polypeptide. Journal of Virology, 29 (1). pp. 352-360. ISSN 0022-538X. PMCID PMC353130. http://resolver.caltech.edu/CaltechAUTHORS:20120725-134020118
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A poliovirus-specific polyuridylic acid [poly(U)] polymerase that copies a polyadenylic acid template complexed to an oligouridylic acid primer was isolated from the membrane fraction of infected HeLa cells and was found to sediment at 4 to 5S on a linear 5 to 20% glycerol gradient. When the poly(U) polymerase was isolated from cells labeled with [^(35)S]methionine and was analyzed by glycerol gradient centrifugation and polyacrylamide gel electrophoresis, the position of only one viral protein was found to correlate with the location of enzyme activity. This protein had an apparent molecular weight of 62,500 based on its electrophoretic mobility relative to that of several molecular weight standards and was designated p63. When the poly(U) polymerase was isolated from the soluble fraction of a cytoplasmic extract, the activity was found to sediment at about 7S. In this case, however, both p63 and NCVP2 (77,000-dalton precursor of p63) cosedimented with the 7S activity peak. When the 7S polymerase activity was purified by phosphocellulose chromatography, both p63 and NCVP2 were found to co-chromatograph with poly(U) polymerase activity. The poliovirus replicase complexed with its endogenous RNA template was isolated from infected cells labeled with [^(35)S]methionine and was centrifuged through a linear 15 to 30% glycerol gradient. The major viral polypeptide component in a 26S peak of replicase activity was p63, but small amounts of other poliovirus proteins were also present. When the replicase-template complex was treated with RNase T1 before centrifugation, a single peak of activity was found that sedimented at 20S and contained only labeled p63. Thus, p63 was found to be the only viral polypeptide in the replicase bound to its endogenous RNA template, and appears to be active as a poly(U) polymerase either as a monomer protein or as a 7S complex.
|Additional Information:||© 1979 American Society for Microbiology. Received for publication 5 July 1978. We thank Victor Ambros, John Rose, Ralf Pettersson, Kahan Leong, Timothy Harrison, and Anthony Shields for their helpful suggestions and for supplying certain materials used in this study. This work was supported by Public Health Service grant AI 08388 from the National Institute of Allergy and Infectious Diseases and grants CA 14051 and CA 12174 from the National Cancer Institute. J.B.F. is the recipient of Public Health Service postdoctoral fellowship F32 AI 05179 from the National Institute of Allergy and Infectious Diseases. D.B. is a Research Professor of the American Cancer Society.|
|PubMed Central ID:||PMC353130|
|Official Citation:||Poliovirus Polyuridylic Acid Polymerase and RNA Replicase Have the Same Viral Polypeptide James B. Flanegan and David Baltimore J. Virol. January 1979 29:352-360|
|Usage Policy:||No commercial reproduction, distribution, display or performance rights in this work are provided.|
|Deposited By:||Ruth Sustaita|
|Deposited On:||25 Jul 2012 22:02|
|Last Modified:||05 Apr 2017 04:38|
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