Identification of compounds that interact with DNA gyrase from Mycobacterium tuberculosis through fragment screening

Abstract

Tuberculosis (TB) is an infectious disease caused by Mycobacterium tuberculosis that affects, approximately, 10 million people a year, and is one of the main causes of death by infections. The treatment of tuberculosis requires the use of at least three drugs that are relatively toxic to patients. As the ideal treatment regimen is approximately six months, individuals are likely to discontinue therapy, which can lead to the appearance of TB drug-resistant strains. Currently, multidrug resistance is a major concern due to the extensive number of strains identified worldwide that are resistant to different molecules used in antituberculosis therapy. In this context, fluoroquinolones are a class of drugs used for the treatment of MDR/RR-TB (multidrug-resistant or rifampicin-resistant TB), whose mechanism of action is the inhibition of DNA gyrase activity, which acts as topoisomerase II that is capable of introducing negative supercoils into DNA, and therefore inhibiting bacterial DNA replication. However, since fluoroquinolones interact with DNA gyrase through one amino acid in the water-metal ion bridge, the enzyme can undergo mutations that interrupt the drug-enzyme interaction, limiting its use in strains resistant to fluoroquinolones. Although in the last 10 years, two new molecules have been approved for use in the treatment of tuberculosis, there is still a need to develop new therapeutic strategies. This project aims to identify small molecules (fragments) that interact with M. tuberculosis DNA gyrase through fragment screening. To do so, expression and purification tests will be done using a construct with subunits GyrA and GyrB fused, as already established in the literature. This construction will be used to screen a library of fragments and the molecules will be further validated by other biophysical techniques. Thus, we expect to identify new chemical scaffolds that can be used in the generation of modifications in existing fluoroquinolones or in the construction of new ligands for this enzyme.