Conversation 11: No. 1
Volume: Conversation 11
Issue Number 1
Protein - TAR RNA Complex and the Design of an Inhibitor of the Tat/TAR RNA Interaction
HIV-1 Tat protein (trans-acting transcriptional activator) regulates transcription in the virus upon binding to the transactivating responsive (TAR) RNA sequence, which is located in the HIV-1 long terminal repeat. Tat is a small protein of 86 residues. Mutation data, binding studies and the use of chemical probes have shown that specific binding occurs between the region centered on a U-rich bulge of TAR RNA and the arginine rich basic region ( residues 49-57) of Tat . The fact that Tat protein plays a critical role in HIV-1 replication suggests that small compounds that disrupt the Tat - TAR interaction will inhibit viral replication. In order to design such a compound, the first step is to model the Tat- TAR complex. The NMR structure of a 29-mer of TAR RNA that contains the minimal RNA region recognised by the Tat protein is known. However, the structure of the basic region of Tat is not well established. We have, therefore carried out a conformational analysis of the 9-residue long basic region that has the sequence RKKRRQRRR. The results indicate that this region has an extended structure. This result is consistent with CD spectroscopic data. We have also carried out a conformational analysis of six synthetic 9-residue peptides whose transactivation abilities have been measured. A superposition of the energy minimized structures of the synthetic peptides with the 9 - residue Tat peptide shows that the r.m.s. deviations of the residues located in the positions 5-8 of the highly active peptides from the corresponding region in Tat are about 0.8Å, whereas the r.m.s. deviations are higher (about 1.5Å) in the case of the peptides with low activities. Thus the conformation of the tetrapeptides is found to correlate well with activity data. This conclusion is supported by molecular modelling of the complex of the 9-residue Tat with the averaged NMR structure of the 29-mer TAR RNA. It is observed that the maximum number of interactions of the Tat peptide with the recognition site on TAR are with residues 5-8 (RQRR) of the Tat peptide. Based on these observations and using the folding rules for dehydro-alanine (D-Ala) derived from our earlier studies, we have designed a tetrapeptide with a sequence Arg- DAla - DAla - Arg. Energy minimization and molecular modelling of the interaction of the designed tetrapeptide with the TAR recogniton site indicate that the tetrapeptide not only has interactions with the recognition site on TAR but also has interactions with additional bases of TAR in the upper stem region as compared to those with Tat. We therefore expect the designed peptide to be an effective inhibitor of the Tat/TAR RNA interaction.
Gita Subba Rao
Department of Biophysics
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