THE STRUCTURE OF AN RNA PSEUDOKNOT THAT INHIBITS HIV-1 REVERSE TRANSCRIPTASE. Craig E. Kundrot, Cindy L. Barnes, and Susan E. Lietzke. Department of Chemistry and Biochemistry, University of Colorado, Boulder, CO 80309-0215.

An RNA pseudoknot inhibitor of HIV-1 reverse transcriptase was isolated by Tuerk, et al., using the in vitro selection technique SELEX. This 26 nucleotide RNA pseudoknot, PK26, specifically inhibits HIV reverse transcriptase with nanomolar affinity. PK26 and three bromo-uridine substituted derivatives were produced by chemical synthesis and crystallized from ammonium acetate. Two of the derivative crystals were suitable for data collection. Diffraction data were collected on cryocooled crystals stabilized with MPD. Native crystals diffracted to 2.9 Å and the derivatives to 3.0 Å. The crystals belong to space group P4_3/122 and have unit cell dimensions a = b = 61.6 Å, c = 98.9 Å. The bromine atoms in the two derivatives were located by Patterson methods and difference Fourier maps and used to produce an initial electron density map to 4 Å resolution (figure of merit = 0.49). The initial map clearly shows that the PK26 molecules coaxially stack in a head-to-head and tail-to-tail orientation. The structure of PK26 will be described and analyzed in terms of extensive chemical modification and nucleotide substitution data available for free PK26 and PK26-HIV RT. Since PK26 also conforms to the pseudoknot motif that promotes ribosome frame shifting, the structure will also be analyzed in terms of potential frame shifting mechanisms.