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Journal of Virology, April 1999, p. 2633-2640, Vol. 73, No. 4
0022-538X/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Isolation from Tobacco Mosaic Virus-Infected Tobacco of a Solubilized Template-Specific RNA-Dependent RNA Polymerase Containing a 126K/183K Protein Heterodimer

Takato Watanabe,1,2 Ayae Honda,1 Akira Iwata,3 Susumu Ueda,3 Tadaaki Hibi,2 and Akira Ishihama1,*

Department of Molecular Genetics, National Institute of Genetics, Mishima, Shizuoka 411-8540,1 Department of Agricultural and Environmental Biology, The University of Tokyo, Yayoi, Bunkyo-ku, Tokyo 113-8657,2 and Nippon Institute for Biological Science, Ohme, Tokyo 198-0024,3 Japan

Received 19 October 1998/Accepted 16 December 1998

The complete nucleotide sequence was determined for the putative RNA polymerase (183K protein) gene of tobacco mosaic virus (TMV) OM strain, which differed from the related strain, vulgare, by 51 positions in its nucleotide sequence and 6 residues in its amino acid sequence. Three segments of this 183K protein, each containing the sequence motif of methyltransferase (M), helicase (H), or RNA-dependent RNA polymerase (P), were expressed in Escherichia coli as fusion proteins with hexahistidine tags, and domain-specific antibodies were raised against purified His-tagged M and P polypeptides. By immunoaffinity purification, a template-specific RNA-dependent RNA polymerase containing a heterodimer of the full-length 183K and 126K (an amino-terminal-proximal portion of the 183K protein) viral proteins was isolated. We propose that the TMV RNA polymerase for minus-strand RNA synthesis is composed of one molecule each of the 183- and 126-kDa proteins, possibly together with two or more host proteins.

* Corresponding author. Mailing address: Department of Molecular Genetics, National Institute of Genetics, Mishima, Shizuoka 411-8540, Japan. Phone: 81-559-81-6741. Fax: 81-559-81-6746. E-mail: aishiham{at}lab.nig.ac.jp.

Journal of Virology, April 1999, p. 2633-2640, Vol. 73, No. 4
0022-538X/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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