Crystal structures, metal activation, and DNA-binding properties of two-domain IdeR from Mycobacterium tuberculosis

Biochemistry. 2007 Jan 16;46(2):436-47. doi: 10.1021/bi0609826.


The iron-dependent regulator IdeR is a key transcriptional regulator of iron uptake in Mycobacterium tuberculosis. In order to increase our insight into the role of the SH3-like third domain of this essential regulator, the metal-binding and DNA-binding properties of two-domain IdeR (2D-IdeR) whose SH3-like domain has been truncated were characterized. The equilibrium dissociation constants for Co2+ and Ni2+ activation of 2D-IdeR for binding to the fxbA operator and the DNA-binding affinities of 2D-IdeR in the presence of excess metal ions were estimated using fluorescence spectroscopy. 2D-IdeR binds to fxbA operator DNA with similar affinity as full-length IdeR in the presence of excess metal ion. However, the Ni2+ concentrations required to activate 2D-IdeR for DNA binding appear to be smaller than that for full-length IdeR while the concentration of Co2+ required for activation remains the same. We have determined the crystal structures of Ni2+-activated 2D-IdeR at 1.96 A resolution and its double dimer complex with the mbtA-mbtB operator DNA in two crystal forms at 2.4 A and 2.6 A, the highest resolutions for DNA complexes for any structures of iron-dependent regulator family members so far. The 2D-IdeR-DNA complex structures confirm the specificity of Ser37 and Pro39 for thymine bases and suggest preferential contacts of Gln43 to cytosine bases of the DNA. In addition, our 2D-IdeR structures reveal a remarkable property of the TEV cleavage sequence remaining after removal of the C-terminal His6. This C-terminal tail promotes crystal contacts by forming a beta-sheet with the corresponding tail of neighboring subunits in two unrelated structures of 2D-IdeR, one with and one without DNA. The contact-promoting properties of this C-terminal TEV cleavage sequence may be beneficial for crystallizing other proteins.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Bacterial Proteins / chemistry*
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism*
  • Base Sequence
  • Binding Sites
  • Crystallography, X-Ray
  • DNA, Bacterial / genetics
  • DNA, Bacterial / metabolism
  • Dimerization
  • Kinetics
  • Macromolecular Substances
  • Metals / metabolism
  • Models, Molecular
  • Mycobacterium tuberculosis / genetics
  • Mycobacterium tuberculosis / metabolism*
  • Nucleic Acid Conformation
  • Protein Structure, Quaternary
  • Protein Structure, Tertiary
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Repressor Proteins / chemistry*
  • Repressor Proteins / genetics
  • Repressor Proteins / metabolism*


  • Bacterial Proteins
  • DNA, Bacterial
  • IdeR protein, Mycobacterium tuberculosis
  • Macromolecular Substances
  • Metals
  • Recombinant Proteins
  • Repressor Proteins

Associated data

  • PDB/2ISY
  • PDB/2ISZ
  • PDB/2IT0