NMR structure of stem-loop SL2 of the HIV-1 Psi RNA packaging signal reveals a novel A-U-A base-triple platform

The genome of the human immunodeficiency virus type-1 (HIV-1) contains a st retch of similar to 120 nucleotides known as the Psi-site that is essential for RNA packaging during virus assembly. These nucleotides have been propo sed to form four stem-loops (SL1-SL4) that have both independent and overla pping functions. Stem-loop SL2 is important for efficient recognition and p ackaging of the full-length, unspliced viral genome, and also contains the major splice-donor site (SD) for mRNA splicing. We have determined the stru cture of the 19-residue SL2 oligoribonucleotide by heteronuclear NMR method s. The structure is generally consistent with the most recent of two earlie r secondary structure predictions, with residues G1-G2-C3-G4 and C6-U7 form ing standard Watson Crick base-pairs with self-complementary residues C16-G 17-C18-C19 and A12-G13, respectively. However, residue A15, which is locate d near the center of the stem, does not form a predicted bulge, and residue s A5 and U14 do not form an expected Watson-Crick base-pair. Instead, these residues form a novel A5-U14-A15 base-triple that appears to be stabilized by hydrogen bonds from A15-H61 and -H62 to A5-N1 and U14-O2, respectively; from A5-H61 to U14-O2, and from C16-H42 to U14-O2'. A kink in the backbone allows the aromatic rings of the sequential U14-A15 residues to be approxi mately co-planar, adopting a stable "platform motif" that is structurally s imilar to the A-A (adenosine) platforms observed in the P4-P6 ribozyme doma in of the Tetrahymena group I intron. Platform motifs generally function in RNA by mediating long-range interactions, and it is therefore possible tha t the A-U-A base-triple platform mediates long-range interactions that eith er stabilize the Psi-RNA or facilitate splicing and/or packaging. Residue G 8 of the G8-G9-U10-G11 tetraloop is stacked above the U7-A12 base-pair, and the remaining tetraloop residues are disordered and available for potentia l interactions with either other RNA or protein components. (C) 2000 Academ ic Press.