The Conformations of the Manganese Transport Regulator of Bacillus subtilis in its Metal-free State

Mark A. DeWitt, Joseph I. Kliegman, John D. Helmann, Richard G. Brennan, David Farrens, Arthur Glasfeld

Research output: Contribution to journalArticle

21 Citations (Scopus)

Abstract

The manganese transport regulator (MntR) from Bacillus subtilis binds cognate DNA sequences in response to elevated manganese concentrations. MntR functions as a homodimer that binds two manganese ions per subunit. Metal binding takes place at the interface of the two domains that comprise each MntR subunit: an N-terminal DNA-binding domain and a C-terminal dimerization domain. In order to elucidate the link between metal binding and activation, a crystallographic study of MntR in its metal-free state has been undertaken. Here we describe the structures of the native protein and a selenomethionine-containing variant, solved to 2.8 Å. The two structures contain five crystallographically unique subunits of MntR, providing diverse views of the metal-free protein. In apo-MntR, as in the manganese complex, the dimer is formed by dyad-related C-terminal domains that provide a conserved structural core. Similarly, each DNA-binding domain largely retains the folded conformation found in metal bound forms of MntR. However, compared to metal-activated MntR, the DNA-binding domains move substantially with respect to the dimer interface in apo-MntR. Overlays of multiple apo-MntR structures indicate that there is a greater range of positioning allowed between N and C-terminal domains in the metal-free state and that the DNA-binding domains of the dimer are farther apart than in the activated complex. To further investigate the conformation of the DNA-binding domain of apo-MntR, a site-directed spin labeling experiment was performed on a mutant of MntR containing cysteine at residue 6. Consistent with the crystallographic results, EPR spectra of the spin-labeled mutant indicate that tertiary structure is conserved in the presence or absence of bound metals, though slightly greater flexibility is present in inactive forms of MntR.

Original languageEnglish (US)
Pages (from-to)1257-1265
Number of pages9
JournalJournal of Molecular Biology
Volume365
Issue number5
DOIs
StatePublished - Feb 2 2007

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Manganese
Bacillus subtilis
Metals
DNA
Selenomethionine
Nucleic Acid Conformation
Dimerization
Cysteine

Keywords

  • allostery
  • metal homeostasis
  • transcriptional regulator
  • X-ray structure

ASJC Scopus subject areas

  • Virology

Cite this

The Conformations of the Manganese Transport Regulator of Bacillus subtilis in its Metal-free State. / DeWitt, Mark A.; Kliegman, Joseph I.; Helmann, John D.; Brennan, Richard G.; Farrens, David; Glasfeld, Arthur.

In: Journal of Molecular Biology, Vol. 365, No. 5, 02.02.2007, p. 1257-1265.

Research output: Contribution to journalArticle

DeWitt, Mark A. ; Kliegman, Joseph I. ; Helmann, John D. ; Brennan, Richard G. ; Farrens, David ; Glasfeld, Arthur. / The Conformations of the Manganese Transport Regulator of Bacillus subtilis in its Metal-free State. In: Journal of Molecular Biology. 2007 ; Vol. 365, No. 5. pp. 1257-1265.
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