Mycobacterial enzymes as anti-tuberculosis drug targets and for use in selective protein modification

• Main Author: Sharma Karkwal, Shalu
• Published: University of Nottingham 2011
• Subjects: 616.995
• Online Access: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.588075

Tuberculosis is an infectious disease caused by Mycobacterium tuberculosis. The mycobacterial cell wall is a characteristic component of mycobacteria and the enzymes involved in its biosynthesis are validated drug targets. Enoyl-ACP reductase (InhA) is involved in the elongation of mycolic acids. Isoniazid (INH), an important anti-tuberculosis drug, is a prodrug and requires activation by a catalase-peroxidase KatG. Our aim was to design inhibitors that can bypass the activation step and hence resistance due to mutations in KatG. In this study recombinant InhA and known INH resistant mutants were cloned, expressed and purified for use in in vitro and cell assays and crystallization studies. Aryl sulfonates tested for inhibitory activity against InhA did not show any promising activity. The galactofuranosyl and arabinofuranosyl transferases GIfT! and EmbA are validated drug targets but lack of structural and mechanistic information of these enzymes has hindered the discovery of novel inhibitors. The structure prediction studies of the enzyme GIITl were conducted. GIITl was also successfully expressed and purified for use in biophysical characterization and crystallization trials. The structure prediction and biophysical characterization studies suggest that GlITl has a typical GT -A a-B fold with a nucleotide binding domain present at the C-terminus. The sequence analysis confirmed that the protein is not membrane bound, while it may be membrane associated. The availability of soluble G1fTl in high yields will facilitate further structural characterization studies.