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Journal of Biochemistry Advance Access originally published online on November 26, 2007
Journal of Biochemistry 2008 143(3):339-347; doi:10.1093/jb/mvm225
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© 2007 The Japanese Biochemical Society.

Recognition of a Bulged RNA by Peptides Derived from the Influenza NS1 Protein

Tatsuhiko Someya*,{dagger}, Kazumi Hosono{dagger}, Kaori Morimura, Hiroshi Takaku and Gota Kawai{ddagger}

Department of Life and Environmental Sciences, Faculty of Engineering, Chiba Institute of Technology, 2-17-1 Tsudanuma, Narashino-shi, Chiba 275-0016, Japan

{ddagger}To whom correspondence should be addressed. Fax: +81-47-478-0425, E-mail: gkawai{at}sea.it-chiba.ac.jp

Received August 27, 2007; Accepted November 12, 2007


  Abstract

A competition assay for RNA binding by the influenza virus NS1 protein using model RNAs, U6–45, corresponding to U6 snRNA revealed that deletion of each of the three bulged-out parts reduced the NS1 protein binding and, in contrast, by deleting all three of the bulged-out parts, simultaneously, and thus producing a double-stranded RNA, the binding was recovered. A common feature of target RNAs of the NS1 protein, U6 snRNA, poly(A) and viral RNA, is the stretch of ‘bulged-out’ A residues. Thus, the NS1 protein was found to recognize either the stretch of ‘bulged-out’ A residues or dsRNA which is also a target of the NS1 protein. Furthermore, a basic peptide, NS1–2, derived from the helix-2 of the RNA binding site of NS1 protein was designed and its binding to the U6 snRNA was analysed by using a model RNA for U6 snRNA, U6–34. The NMR signals due to H8/H6 and H1' of U6–34 were assigned and their changes upon binding of NS1–2 were analysed. It was indicated that NS1–2 interacts with the residues in the bulge-out region of U6–34. These results suggest that NS1–2 recognizes the U6 snRNA in a similar manner to NS1 protein.

Key Words: gel shift, influenza virus, NMR, NS1 protein, RNA-binding protein, U6 snRNA

Abbreviations: dsRNA, double-stranded RNA; PAGE, polyacrylamide gel electrophoresis; NS1, non-structural protein 1; snRNA, small nuclear RNA

*Present address: Graduate School of Life and Environmental Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba-shi, Ibaraki 305-8572, Japan.

{dagger}These authors contribute to this work equally.


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