Pseudoknots: RNA structures with diverse functions.
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Base Composition↗
Biomedical subjects
David W Staple
Publications and source records attributed to David W Staple.
Solution structure and thermodynamic investigation of the HIV-1 frameshift inducing element.
Expression of the HIV reverse transcriptase and other essential viral enzymes requires a -1 translational frameshift. The frameshift event is induced by two highly conserved RNA elements within the HIV-1 mRNA: a UUUUUUA heptamer known as the slippery sequence, and a downstream RNA structure. Here, we report structural and thermodynamic evidence that the HIV-1 frameshift site RNA forms a stem-loop and lower helix separated by a three-purine bulge. We have determined the structure of the 45 nucleotide frameshift site RNA using multidimensional heteronuclear nuclear magnetic resonance (NMR) methods. The upper helix is highly thermostable (T(m)>90 degrees C), forming 11 Watson-Crick base-pairs capped by a stable ACAA tetraloop. The eight base-pair lower helix was found to be only moderately stable (T(m)=47 degrees C). A three-purine bulge separates the highly stable upper helix from the lower helix. Base stacking in the bulge forms a wedge, introducing a 60 degrees bend between the helices. Interestingly, this bend is similar to those seen in a number of frameshift inducing pseudoknots for which structures have been solved. The lower helix must denature to allow the ribosome access to the slippery site, but likely functions as a positioning element that enhances frameshift efficiency.
Frameshifting, Ribosomal↗
Solution structure of the HIV-1 frameshift inducing stem-loop RNA.
The translation of reverse transcriptase and other essential viral proteins from the HIV-1 Pol mRNA requires a programmed -1 ribosomal frameshift. This frameshift is induced by two highly conserved elements within the HIV-1 mRNA: a slippery sequence comprised of a UUUUUUA heptamer, and a downstream stem-loop structure. We have determined the structure of the HIV-1 frameshift inducing RNA stem-loop, using multidimensional heteronuclear nuclear magnetic resonance (NMR) methods. The 22 nucleotide RNA solution structure [root mean squared deviation (r.m.s.d.) = 1.2 A] was determined from 475 nuclear Overhauser effect (NOE)-derived distance restrains, 20 residual dipolar couplings and direct detection of hydrogen bonds via scalar couplings. We find that the frameshift inducing stem-loop is an A-form helix capped by a structured ACAA tetraloop. The ACAA tetraloop is stabilized by an equilateral 5' and 3' stacking pattern, a sheared A-A pair and a cross-strand hydrogen bond. Unexpectedly, the ACAA tetraloop structure is nearly identical to a known tetraloop fold, previously identified in the RNase III recognition site from Saccharomyces cerevisiae.
Adenine↗