Mapping binding sites of nucleic acid ligands targeting unique structural features of the HIV-1 polypurine tract by tandem mass spectrometry

摘要

Initiation of reverse transcription in HIV-1 requires the accurate selection of a distinctive polypurine tract (PPT) from within a considerably larger RNA/DNA hybrid (Fig. 1). At a later stage, PPT must be excised from (+) DNA with equivalent precision. Therefore, PPT selection and removal imply a high level of selectivity by reverse transcriptase (RT), which must recognize very specific structural characteristics of this purine-rich hybrid. A variety of biochemical and structural studies indicate that two regions of the HIV-1 PPT-containing RNA/DNA hybrid deviate from standard Watson-Crick geometry. We have employed electrospray ionization (ESI) and Fourier transform ion cyclotron resonance (FTICR) mass spectrometry to investigate the non-covalent interactions between PPT and small ligands that could specifically recognize these unique structural features. The observed sites were mapped directly by tandem mass spectrometry, in an effort to understand the binding determinants and obtain valuable information for the development of new inhibitors aimed at interfering with normal RT activity.