RNA aptamer inhibitors of HIV reverse transcriptase: Molecular evolution and determinants of target specificity

The nature of intermolecular interactions between proteins and nucleic acids is governed by evolutionary constraints imposed on those biological macromolecules. With the development of in vitro selection techniques for nucleic acid aptamers, we have been provided a powerful tool for use in our quest to better understand the influences of molecular evolution on the determinants of target specificity between nucleic acids and proteins. RNA aptamers selected for binding to the reverse transcriptase (RT) of type I human immunodeficiency virus (HIV-1) inhibit the enzyme's activity by competing for primer-template recognition. When expressed in cultured lymphocytes, aptamers protect the cells from viral replication by inhibiting viral genome replication. Two prominent questions follow from these results: (1) Do aptamers preferentially inhibit viral genome replication at specific points during the multi-step process? and (2) What is the breadth of cross-clade efficacy of RT aptamers? I explored these details of aptamer potency against a panel of recombinant HIV-1, HIV-2, and SIVcpz RTs using a series of assays designed to mimic specific steps of viral genome replication. My findings reveal variability in aptamer potency against discrete steps of reverse transcription, as well as dramatic differences in aptamer potency across viral clades. These results can be understood in part as a function of aptamer structural variability and a single amino acid polymorphism in RT within viral populations. This same principle is evidenced by a project that had nothing to do with HIV-1, but everything to do with the evolution of molecular recognition. Specifically, analysis of the evolutionary pathways between aptamers selected for binding to the nucleotide cofactor FAD and mononucleotide GMP reveals that functional sequence space is very plastic even between less-closely related targets for aptamer binding. Overall, the results from these two projects suggest that despite readily available evolutionary pathways by which HIV could achieve resistance to certain aptamers, broad-spectrum aptamer therapeutics may still be available.