http://www.abbs.info E-mail: [email protected]
ISSN 0582-9879 Acta Biochimica et Biophysica Sinica 2004, 36(7): 457–466 CN 31-1300/Q
Nucleic Acid Binding Activity of Pns6 Encoded by Genome Segment 6 of Rice Ragged Stunt Oryzavirus
Chao-Gang SHAO#, Hui-Juan LÜ#, Jian-Hua WU, and Zu-Xun GONG*
Key Laboratory of Proteomics, Institute of Biochemistry
and Cell Biology, Shanghai Institutes of Life Sciences, Chinese Academy of
Sciences,Graduate School of the Chinese Academy of Sciences, Shanghai 200031,
China
Abstract The ORF of genome segment 6 (S6) of rice ragged stunt oryzavirus (RRSV) Philippines isolate was cloned and sequenced based on the S6 sequence of the Thailand isolate. Pns6, the 71 kD product of S6 expressed in E. coli, was demonstrated to be a viral non-structural protein of RRSV by Western blotting. The gel mobility shift assays showed that Pns6 had nucleic acid binding activity. Pns6 could interact with single- and double-stranded forms of DNA and RNA, showing a preference for single-stranded nucleic acid and a slight preference for RRSV ssRNA over the rice ssRNA, as demonstrated by both competition and displacement assays. The binding of Pns6 to nucleic acids is strong and sequence non-specific. By using five truncated derivatives of Pns6, it was found that the basic region from amino acid 201 to 273 of Pns6 was the unique nucleic acid binding domain. Subcellular fractionation of leaf tissues of RRSV-infected rice plants and subsequent Western blotting had shown that Pns6 accumulated predominately in the cytoplasmic membrane fraction. The possible role of RRSV Pns6 in virus replication and assembly is discussed.
Key words rice ragged stunt oryzavirus (RRSV); nucleic acid binding; non-structural protein; S6
-----------------
Received: March 3, 2004 Accepted: May 28, 2004
This work was supported by the grants from Rockefeller Foundation, the AusAid under its Australia-China Institutional Links Program, and the National Natural Science Foundation of China (No. 30170042)
#These two authors contributed equally to this work
*Corresponding author: Tel, 86-21-54921220; Fax, 86-21-54921225;
E-mail, [email protected]