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A loop 2 cytidine-stem 1 minor groove interaction as a positive determinant for pseudoknot-stimulated –1 ribosomal frameshifting
*Department of Biochemistry and Biophysics, 2128 TAMU, Texas A&M University, College Station, TX 77843-2128; and Department of Molecular Biology, The Scripps Research Institute, MB33, 10550 North Torrey Pines Road, La Jolla, CA 92037
Communicated by Ignacio Tinoco, Jr., University of California, Berkeley, CA, July 20, 2005 (received for review January 9, 2005)
The molecular determinants of stimulation of –1 programmed ribosomal frameshifting (–1 PRF) by RNA pseudoknots are poorly understood. Sugarcane yellow leaf virus (ScYLV) encodes a 28-nt mRNA pseudoknot that promotes –1 PRF between the P1 (protease) and P2 (polymerase) genes in plant luteoviruses. The solution structure of the ScYLV pseudoknot reveals a well ordered loop 2 (L2) that exhibits continuous stacking of A20 through C27 in the minor groove of the upper stem 1 (S1), with C25 flipped out of the triple-stranded stack. Five consecutive triple base pairs flank the helical junction where the 3' nucleotide of L2, C27, adopts a cytidine 27 N3-cytidine 14 2'-OH hydrogen bonding interaction with the C14-G7 base pair. This interaction is isosteric with the adenosine N1–2'-OH interaction in the related mRNA from beet western yellows virus (BWYV); however, the ScYLV and BWYV mRNA structures differ in their detailed L2–S1 hydrogen bonding and L2 stacking interactions. Functional analyses of ScYLV/BWYV chimeric pseudoknots reveal that the ScYLV RNA stimulates a higher level of –1 PRF (15 ± 2%) relative to the BWYV pseudoknot (6 ± 1%), a difference traced largely to the identity of the 3' nucleotide of L2 (C27 vs. A25 in BWYV). Strikingly, C27A ScYLV RNA is a poor frameshift stimulator (2.0%) and is destabilized by 
1.5 kcal·mol–1 (pH 7.0, 37°C) with respect to the wild-type pseudoknot. These studies establish that the precise network of weak interactions nearest the helical junction in structurally similar pseudoknots make an important contribution to setting the frameshift efficiency in mRNAs.
base triple | RNA pseudoknot | translational recoding
Abbreviations: BWYV, beet western yellows virus; L1, loop 1; L2, loop 2; –1 PRF, –1 programmed ribosomal frameshifting; RDC, residual dipolar coupling; S1, stem 1; S2, stem 2; ScYLV, sugarcane yellow leaf virus.
Data deposition: The atomic coordinates have been deposited in the Protein Data Bank, www.pdb.org [PDB ID codes 1YG3
© 2005 by The National Academy of Sciences of the USA
This article has been cited by other articles in HighWire Press-hosted journals:
To whom correspondence should be addressed. E-mail: giedroc{at}tamu.edu. 
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