Abstract
The virion-associated RNA polymerase and the structural nucleoprotein of influenza A virus were separated by sodium dodecyl sulfate/PAGE, electroblotted to a polyvinylidine membrane, and eluted with good recovery from the membrane. After renaturation by incubating with Escherichia coli thioredoxin, these proteins were active in a reconstituted in vitro transcription reaction with purified genomic RNAs. All four proteins (i.e., the three subunits of the RNA polymerase as well as the structural nucleoprotein) were required for activity. The RNA products were plus-strand, mRNA-sized species.
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Selected References
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- Aboud M., Hassan Y. Accumulation and breakdown of RNA-deficient intracellular virus particles in interferon-treated NIH 3T3 cells chronically producing Moloney murine leukemia virus. J Virol. 1983 Feb;45(2):489–495. doi: 10.1128/jvi.45.2.489-495.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Akkina R. K., Chambers T. M., Londo D. R., Nayak D. P. Intracellular localization of the viral polymerase proteins in cells infected with influenza virus and cells expressing PB1 protein from cloned cDNA. J Virol. 1987 Jul;61(7):2217–2224. doi: 10.1128/jvi.61.7.2217-2224.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bailey J. M., Davidson N. Methylmercury as a reversible denaturing agent for agarose gel electrophoresis. Anal Biochem. 1976 Jan;70(1):75–85. doi: 10.1016/s0003-2697(76)80049-8. [DOI] [PubMed] [Google Scholar]
- Blaas D., Patzelt E., Kuechler E. Identification of the cap binding protein of influenza virus. Nucleic Acids Res. 1982 Aug 11;10(15):4803–4812. doi: 10.1093/nar/10.15.4803. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bouloy M., Plotch S. J., Krug R. M. Globin mRNAs are primers for the transcription of influenza viral RNA in vitro. Proc Natl Acad Sci U S A. 1978 Oct;75(10):4886–4890. doi: 10.1073/pnas.75.10.4886. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Braam J., Ulmanen I., Krug R. M. Molecular model of a eucaryotic transcription complex: functions and movements of influenza P proteins during capped RNA-primed transcription. Cell. 1983 Sep;34(2):609–618. doi: 10.1016/0092-8674(83)90393-8. [DOI] [PubMed] [Google Scholar]
- Detjen B. M., St Angelo C., Katze M. G., Krug R. M. The three influenza virus polymerase (P) proteins not associated with viral nucleocapsids in the infected cell are in the form of a complex. J Virol. 1987 Jan;61(1):16–22. doi: 10.1128/jvi.61.1.16-22.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hay A. J., Skehel J. J., McCauley J. Characterization of influenza virus RNA complete transcripts. Virology. 1982 Jan 30;116(2):517–522. doi: 10.1016/0042-6822(82)90144-1. [DOI] [PubMed] [Google Scholar]
- Horisberger M. A. The large P proteins of influenza A viruses are composed of one acidic and two basic polypeptides. Virology. 1980 Nov;107(1):302–305. doi: 10.1016/0042-6822(80)90296-2. [DOI] [PubMed] [Google Scholar]
- Jones I. M., Reay P. A., Philpott K. L. Nuclear location of all three influenza polymerase proteins and a nuclear signal in polymerase PB2. EMBO J. 1986 Sep;5(9):2371–2376. doi: 10.1002/j.1460-2075.1986.tb04506.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kato A., Mizumoto K., Ishihama A. Purification and enzymatic properties of an RNA polymerase-RNA complex from influenza virus. Virus Res. 1985 Sep;3(2):115–127. doi: 10.1016/0168-1702(85)90002-4. [DOI] [PubMed] [Google Scholar]
- Kingsbury D. W., Jones I. M., Murti K. G. Assembly of influenza ribonucleoprotein in vitro using recombinant nucleoprotein. Virology. 1987 Feb;156(2):396–403. doi: 10.1016/0042-6822(87)90419-3. [DOI] [PubMed] [Google Scholar]
- Krystal M., Li R., Lyles D., Pavlakis G., Palese P. Expression of the three influenza virus polymerase proteins in a single cell allows growth complementation of viral mutants. Proc Natl Acad Sci U S A. 1986 Apr;83(8):2709–2713. doi: 10.1073/pnas.83.8.2709. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lamb R. A., Choppin P. W. The gene structure and replication of influenza virus. Annu Rev Biochem. 1983;52:467–506. doi: 10.1146/annurev.bi.52.070183.002343. [DOI] [PubMed] [Google Scholar]
- Laver W. G. Crystallization and peptide maps of neuraminidase "heads" from H2N2 and H3N2 influenza virus strains. Virology. 1978 May 1;86(1):78–87. doi: 10.1016/0042-6822(78)90009-0. [DOI] [PubMed] [Google Scholar]
- Pigiet V. P., Schuster B. J. Thioredoxin-catalyzed refolding of disulfide-containing proteins. Proc Natl Acad Sci U S A. 1986 Oct;83(20):7643–7647. doi: 10.1073/pnas.83.20.7643. [DOI] [PMC free article] [PubMed] [Google Scholar]
- St Angelo C., Smith G. E., Summers M. D., Krug R. M. Two of the three influenza viral polymerase proteins expressed by using baculovirus vectors form a complex in insect cells. J Virol. 1987 Feb;61(2):361–365. doi: 10.1128/jvi.61.2.361-365.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Szewczyk B., Summers D. F. Preparative elution of proteins blotted to Immobilon membranes. Anal Biochem. 1988 Jan;168(1):48–53. doi: 10.1016/0003-2697(88)90008-5. [DOI] [PubMed] [Google Scholar]
- Ulmanen I., Broni B. A., Krug R. M. Role of two of the influenza virus core P proteins in recognizing cap 1 structures (m7GpppNm) on RNAs and in initiating viral RNA transcription. Proc Natl Acad Sci U S A. 1981 Dec;78(12):7355–7359. doi: 10.1073/pnas.78.12.7355. [DOI] [PMC free article] [PubMed] [Google Scholar]