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Solution structure of Mycobacterium tuberculosis NmtR in the apo state: insights into Ni(II)-mediated allostery.

ABSTRACT: Mycobacterium tuberculosis is an obligate human respiratory pathogen that encodes approximately 10 arsenic repressor (ArsR) family regulatory proteins that allow the organism to respond to a wide range of changes in its immediate microenvironment. How individual ArsR repressors have evolved to respond to selective stimuli is of intrinsic interest. The Ni(II)/Co(II)-specific repressor NmtR and related actinomycete nickel sensors harbor a conserved N-terminal ?-NH(2)-Gly2-His3-Gly4 sequence. Here, we present the solution structure of homodimeric apo-NmtR and show that the core of the molecule adopts a typical winged-helix ArsR repressor (?1-?2-?3-?R-?1-?2-?5) "open conformation" that is similar to that of the related zinc sensor Staphylococcus aureus CzrA, but harboring long, flexible N-terminal (residues 2-16) and C-terminal (residues 109-120) extensions. Binding of Ni(II) to the regulatory sites induces strong paramagnetic broadening of the ?5 helical region and the extreme N-terminal tail to residue 10. Ratiometric pulse chase amidination mass spectrometry reveals that the rate of amidination of the ?-amino group of Gly2 is strongly attenuated in the Ni(II) complex relative to the apo state and noncognate Zn(II) complex. Ni(II) binding also induces dynamic disorder on the microsecond to millisecond time scale of key DNA interacting regions that likely contributes to the negative regulation of DNA binding by Ni(II). Molecular dynamics simulations and quantum chemical calculations reveal that NmtR readily accommodates a distal Ni(II) hexacoordination model involving the ?-amine and His3 of the N-terminal region and ?5 residues Asp91', His93', His104, and His107, which collectively define a new metal sensing site configuration in ArsR family regulators.

SUBMITTER: Lee CW

PROVIDER: S-EPMC3500661 | biostudies-literature | 2012 Mar

REPOSITORIES: biostudies-literature

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Solution structure of Mycobacterium tuberculosis NmtR in the apo state: insights into Ni(II)-mediated allostery.

Lee Chul Won CW Chakravorty Dhruva K DK Chang Feng-Ming James FM Reyes-Caballero Hermes H Ye Yuzhen Y Merz Kenneth M KM Giedroc David P DP

Biochemistry 20120314 12

Mycobacterium tuberculosis is an obligate human respiratory pathogen that encodes approximately 10 arsenic repressor (ArsR) family regulatory proteins that allow the organism to respond to a wide range of changes in its immediate microenvironment. How individual ArsR repressors have evolved to respond to selective stimuli is of intrinsic interest. The Ni(II)/Co(II)-specific repressor NmtR and related actinomycete nickel sensors harbor a conserved N-terminal α-NH(2)-Gly2-His3-Gly4 sequence. Here, ...[more]

PMID: 22394357

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Project description:Combating the spread of drug resistant tuberculosis is a global health priority. Whole genome association studies are being applied to identify genetic determinants of resistance to anti-tuberculosis drugs. Protein structure and interaction modelling are used to understand the functional effects of putative mutations and provide insight into the molecular mechanisms leading to resistance.To investigate the potential utility of these approaches, we analysed the genomes of 144 Mycobacterium tuberculosis clinical isolates from The Special Programme for Research and Training in Tropical Diseases (TDR) collection sourced from 20 countries in four continents. A genome-wide approach was applied to 127 isolates to identify polymorphisms associated with minimum inhibitory concentrations for first-line anti-tuberculosis drugs. In addition, the effect of identified candidate mutations on protein stability and interactions was assessed quantitatively with well-established computational methods.The analysis revealed that mutations in the genes rpoB (rifampicin), katG (isoniazid), inhA-promoter (isoniazid), rpsL (streptomycin) and embB (ethambutol) were responsible for the majority of resistance observed. A subset of the mutations identified in rpoB and katG were predicted to affect protein stability. Further, a strong direct correlation was observed between the minimum inhibitory concentration values and the distance of the mutated residues in the three-dimensional structures of rpoB and katG to their respective drugs binding sites.Using the TDR resource, we demonstrate the usefulness of whole genome association and convergent evolution approaches to detect known and potentially novel mutations associated with drug resistance. Further, protein structural modelling could provide a means of predicting the impact of polymorphisms on drug efficacy in the absence of phenotypic data. These approaches could ultimately lead to novel resistance mutations to improve the design of tuberculosis control measures, such as diagnostics, and inform patient management.

Dataset Information

Solution structure of Mycobacterium tuberculosis NmtR in the apo state: insights into Ni(II)-mediated allostery.

Publications

Solution structure of Mycobacterium tuberculosis NmtR in the apo state: insights into Ni(II)-mediated allostery.

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