Nucleic Acids Research Advance Access published online on September 26, 2009
Nucleic Acids Research, doi:10.1093/nar/gkp793
RNA |
The HSV-1 ICP27 RGG box specifically binds flexible, GC-rich sequences but not G-quartet structures
1Department of Microbiology and Molecular Genetics and 2Department of Molecular Biology and Biochemistry, University of California Irvine, Irvine, CA 92697, USA
*To whom correspondence should be addressed. Tel: +1 949 856 7570; Fax: +1 949 824 8598; Email: rmsandri{at}uci.edu
Correspondence may also be addressed to Melanie J. Cocco. Fax: +1 949 824 8551; Email: mcocco{at}uci.edu
Received June 22, 2009. Revised September 8, 2009. Accepted September 9, 2009.
Herpes simplex virus 1 (HSV-1) protein ICP27, an important regulator for viral gene expression, directly recognizes and exports viral RNA through an N-terminal RGG box RNA binding motif, which is necessary and sufficient for RNA binding. An ICP27 N-terminal peptide, including the RGG box RNA binding motif, was expressed and its binding specificity was analyzed using EMSA and SELEX. DNA oligonucleotides corresponding to HSV-1 glycoprotein C (gC) mRNA, identified in a yeast three-hybrid analysis, were screened for binding to the ICP27 N-terminal peptide in EMSA experiments. The ICP27 N-terminus was able to bind most gC substrates. Notably, the ICP27 RGG box was unable to bind G-quartet structures recognized by the RGG domains of other proteins. SELEX analysis identified GC-rich RNA sequences as a common feature of recognition. NMR analysis of SELEX and gC sequences revealed that sequences able to bind to ICP27 did not form secondary structures and conversely, sequences that were not able to bind to ICP27 gave spectra consistent with base-pairing. Therefore, the ICP27 RGG box is unique in its recognition of nucleic acid sequences compared to other RGG box proteins; it prefers flexible, GC-rich substrates that do not form stable secondary structures.