In vitro selection identifies key determinants for loop-loop interactions: RNA aptamers selective for the TAR RNA element of HIV-1.
We selected RNA aptamers specific for the trans-activation responsive (TAR) RNA, a stem-loop structure crucial for the transcription of the integrated genome of the human immunodeficiency virus. Most of the selected sequences could be folded as imperfect hairpins and displayed a 5'-GUCCCAGA-3' consensus motif constituting the apical loop. The six central bases of this consensus sequence are complementary to the entire TAR loop, leading to the formation of TAR RNA-aptamer "kissing" complexes. The consensus G and A residues closing the aptamer loop contributed to the high affinity (Kd = 30 nM at 23 degrees C) of the aptamers for the TAR RNA. This G A pair was shown to be crucial for binding to TAR at a low magnesium concentration. The selection also identified 5'-PuPy and 5'-PyPu base pairs at alpha and beta positions of the stem, next to the loop, respectively. This strategy offered a way to identify key determinants of loop-loop interactions and to generate high affinity ligands of TAR RNA structure.
Ducongé
F
F
Institut National de la Santé et de la Recherche Médicale U 386, IFR Pathologies Infectieuses, Université Victor Segalen, Bordeaux, France.
Toulmé
J J
JJ
D020029:Base Pairing; D001483:Base Sequence; D001665:Binding Sites; D016384:Consensus Sequence; D017931:DNA Primers; D016325:HIV Long Terminal Repeat / Q000235:genetics*; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009843:Oligoribonucleotides / Q000737:chemistry*; D016133:Polymerase Chain Reaction; D012367:RNA, Viral / Q000737:chemistry* / Q000235:genetics*; D013329:Structure-Activity Relationship
D017931:DNA Primers; D009843:Oligoribonucleotides; D012367:RNA, Viral