Mycobacterium tuberculosis Rv1474c is a TetR-like transcriptional repressor that regulates aconitase, an essential enzyme and RNA-binding protein, in an iron-responsive manner
Kannan Balakrishnan, Krishnaveni Mohareer, Sharmistha Banerjee
Mycobacterium tuberculosis (M.tb), tuberculosis (TB) causing bacteria, employs several mechanisms to maintain iron homeostasis which is critical for its survival and pathogenesis. M.tb aconitase (Acn), a [4Fe-4S] cluster-containing essential protein, apart from participating in energy cycle, also binds to predicted iron-responsive RNA. In this study, we identified Rv1474c as a TetR-like repressor of its operonic partner acn and carried out its biochemical and functional characterization. The binding motif for Rv1474c in the upstream region of acn (Rv1475c)-Rv1474c operon was verified by gel-shift assays. Reporter assays in E. coli followed by over-expression studies in mycobacteria, using both wild type and a DNA-binding defective mutant, demonstrated Rv1474c as a Tet-R like repressor of acn. Rv1474c, beside binding tetracycline, could also bind iron which negatively influenced its DNA binding activity. A consistent decrease in the relative transcript levels of acn when M.tb was grown in iron-deficient conditions as compared to either normal or other stress conditions, indicated regulation of acn by Rv1474c in an iron-responsive manner in vivo. The absence of homologs in the human host and its association with indispensable iron homeostasis makes Rv1474c an attractive target for designing novel anti-mycobacterials.
Circular dichroism, Secondary structure, Ligand binding, Biochemistry