Using a Novel Hybrid Method for the Design of Inhibitors Targeting DnaG Primase of Mycobacterium Tuberculosis

The DNA replication process in Mycobacterium tuberculosis (Mtb) is a promising but underexploited target for the development of novel antibiotics. We have developed an approach to identify inhibitors for Mtb DnaG primase, which is a key enzyme in the DNA replication machinery of Mtb. For the development process we have used DNA primase from bacteriophage T7. T7 DNA primase has several structural features that are similar to bacterial (including Mtb) primases, making it an ideal model to study bacterial primases. Using NMR screening, fragment molecules that bind T7 primase were identified and then exploited in virtual filtration to select larger molecules from a virtual library. The molecules were docked to the primase active site using the available T7 primase crystal structure and ranked based on their binding energies to identify the best candidates for functional and structural investigations. Biochemical assays revealed that some of the molecules inhibit T7 primase-dependent DNA replication. The binding mechanism was delineated via NMR spectroscopy. Importantly, some of the molecules inhibit also the activity of DnaG primase of Mtb. Our studies yielded new class of potential antituberculotics and provided new tools for fragment-based lead discovery