Selection and design of high-affinity RNA ligands for HIV-1 Rev.
We have used in vitro selection to isolate minimal, high-affinity RNA ligands for the Rev protein of HIV-1. Sequence analysis reveals that the tightest binding aptamers exhibit some similarity to a Rev-binding element (RBE) localized within the Rev-responsive element (RRE), but also contain novel sequence and structural motifs. A short helical stem and bulged nucleotides (nt) CUC ... UYGAG that have no counterpart in the wild-type (wt) element contribute to high-affinity binding. We have designed and synthesized a short (37 nt) RNA molecule that incorporates this motif; this RNA ligand has from three- to fivefold tighter binding than the full-length wt element, and up to 16-fold tighter than minimal wt RBEs. A guanosine:guanosine pairing that is postulated to occur in the wt element has been altered to other base pairings in the context of our optimized minimal element. RNAs that contain non-Watson-Crick base pairings, that can be modeled as isosteric to the wt G:G pair, bind Rev up to 160-fold tighter than elements that contain canonical Watson-Crick pairings or non-isosteric mismatches. These results support the hypothesis that Rev recognizes structural features associated with a non-Watson-Crick base pair.
Giver
L
L
Department of Chemistry, Indiana University, Bloomington 47405.
Bartel
D P
DP
Zapp
M L
ML
Green
M R
MR
Ellington
A D
AD
D001482:Base Composition; D001483:Base Sequence; D015694:Gene Products, rev / Q000737:chemistry*; D008024:Ligands; D008956:Models, Chemical; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D012313:RNA / Q000138:chemical synthesis*
D015694:Gene Products, rev; D008024:Ligands; D012313:RNA