Sanjay Sunder

Chemical Trapping of Ternary Complexes of Human Immunodeficiency Virus Type 1 Integrase, Divalent Metal, and DNA Substrates Containing an Abasic Site IMPLICATIONS FOR THE ROLE OF LYSINE 136 IN DNA BINDING

We report a novel assay for monitoring the DNA binding of human immunodeficiency virus type 1 (HIV-1) integrase and the effect of cofactors and inhibitors. The assay uses depurinated oligonucleotides that can form a Schiff base between the aldehydic abasic site and a nearby enzyme lysine ε-amino group which can subsequently be trapped by reduction with sodium borohydride. Chemically depurinated duplex substrates representing the U5 end of the HIV-1 DNA were initially used. We next substituted an enzymatically generated abasic site for each of 10 nucleotides normally present in a 21-mer duplex oligonucleotide representing the U5 end of the HIV-1 DNA. Using HIV-1, HIV-2, or simian immunodeficiency virus integrases, the amount of covalent enzyme-DNA complex trapped decreased as the abasic site was moved away from the conserved CA dinucleotide. The enzyme-DNA complexes formed in the presence of manganese were not reversed by subsequent addition of EDTA, indicating that the divalent metal required for integrase catalysis is tightly bound in a ternary enzyme-metal-DNA complex. Both the N- and C-terminal domains of integrase contributed to efficient DNA binding, and mutation of Lys-136 significantly reduced Schiff base formation, implicating this residue in viral DNA binding.

Design and discovery of HIV-1 integrase inhibitors

The rapid emergence of human immunodeficiency virus (HIV) strains resistant to available drugs implies that effective treatment modalities will require the use of a combination of drugs targeting different sites of the HIV life cycle. Integrase is emerging as a novel target for intervention by chemotherapeutics. As part of a program to develop novel antiviral agents, we have studied a large number of compounds with activity against HIV-1 replication in the National Cancer Institute (NCI) Antiviral Drug Program. In this review, we present a comprehensive review of all such inhibitors reported to date.