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Identification of a copper-binding metallothionein in pathogenic mycobacteria

Nature Chemical Biology volume 4, pages 609–616 (2008)Cite this article

Abstract

A screen of a genomic library from Mycobacterium tuberculosis (Mtb) identified a small, unannotated open reading frame (MT0196) that encodes a 4.9-kDa, cysteine-rich protein. Despite extensive nucleotide divergence, the amino acid sequence is highly conserved among mycobacteria that are pathogenic in vertebrate hosts. We synthesized the protein and found that it preferentially binds up to six Cu(I) ions in a solvent-shielded core. Copper, cadmium and compounds that generate nitric oxide or superoxide induced the gene's expression in Mtb up to 1,000-fold above normal expression. The native protein bound copper within Mtb and partially protected Mtb from copper toxicity. We propose that the product of the MT0196 gene be named mycobacterial metallothionein (MymT). To our knowledge, MymT is the first metallothionein of a Gram-positive bacterium with a demonstrated function.

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Figure 1: Identification of MymT as a polypeptide with MT-like metal-binding motifs.
Figure 2: Mass spectra of MymT complexed with Cu(I) or Zn(II).
Figure 3: MymT binds Cu(I) in a solvent-shielded, luminescent core.
Figure 4: Regulation of mymT transcript abundance.
Figure 5: Generation of a ΔmymT mutant in Mtb.
Figure 6: Sensitivity of the Mtb ΔmymT mutant to cuprous ion.
Figure 7: Generation of Cu(I) by NO.

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Acknowledgements

We thank D. Domaille and C. Chang (University of California, Berkeley); H. Erdjument-Bromage and P. Tempst (Protein Center, Sloan-Kettering Institute); P. Wille, A. Morrishow and S. Gross (Weill Medical College); T. Kapoor and J. Cherian (Rockefeller University); and M. Gurney, J. Singh and R. Samy (deCODE Chemistry) for help with experiments not included here. We thank J. Cox (University of California, San Francisco) and V. Rao and M. Glickman (Sloan Kettering Institute) for phages; N. Chandramouli (Rockefeller University) for synthesis of MymT; S. Ehrt, A. Ding and K. Rhee for critical review of the manuscript; and N. Brot, S. Marras, S. Walters, M. Monteleone and members of the labs of C. Nathan, S. Ehrt and A. Ding for advice. We thank the anonymous reviewers for important guidance. This work was supported by US National Institutes of Health grant AI 62559. The Department of Microbiology and Immunology of Weill Cornell Medical College is supported by the William Randolph Hearst Foundation.

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  1. Program in Immunology and Microbial Pathogenesis and Program in Molecular Biology, Weill Graduate School of Medical Sciences of Cornell University, 1300 York Avenue, New York, New York 10065, USA.

Authors and Affiliations

  1. Department of Microbiology and Immunology, Weill Cornell Medical College, 1300 York Avenue, New York, 10065, New York, USA

    Ben Gold, Ruslana Bryk, Julia Roberts, Xiuju Jiang & Carl Nathan

  2. Proteomics Resource Center, The Rockefeller University, 1230 York Avenue, New York, 10065, New York, USA

    Haiteng Deng

  3. Department of Molecular Genetics, Public Health Research Institute, 225 Warren Street, Newark, 07103, New Jersey, USA

    Diana Vargas

  4. Department of Biochemistry, Weill Cornell Medical College, 1300 York Avenue, New York, 10065, New York, USA

    David Eliezer

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Contributions

B.G. initiated the study, designed and performed experiments, analyzed data and co-wrote the manuscript. H.D. designed, performed and analyzed ESI-MS studies and supervised synthesis of MymT. R.B. and D.E. helped design experiments and analyze data. D.V. designed and performed qRT-PCR experiments and analyzed data. J.R. and X.J. performed experiments. C.N. helped design experiments, analyze data and write the manuscript.

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Correspondence to Carl Nathan.

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Gold, B., Deng, H., Bryk, R. et al. Identification of a copper-binding metallothionein in pathogenic mycobacteria. Nat Chem Biol 4, 609–616 (2008). https://doi.org/10.1038/nchembio.109

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