Structure of the carboxy-terminal domain of Mycobacterium tuberculosis CarD protein: An essential rRNA transcriptional regulator

Shanti P. Gangwar, Sita R. Meena, Ajay K. Saxena

Research output: Contribution to journalArticle

2 Scopus citations

Abstract

The CarD protein is highly expressed in mycobacterial strains under basal conditions and is transcriptionally induced during multiple types of genotoxic stress and starvation. The CarD protein binds the β subunit of RNA polymerase and influences gene expression. The disruption of interactions between CarD and the β subunit of RNA polymerase has a significant effect on mycobacterial survival, resistance to stress and pathogenesis. To understand the structure of CarD and its interaction with the β subunit of RNA polymerase, Mycobacterium tuberculosis CarD (MtbCarD) and the Thermus aquaticus RNA polymerase β subunit were recombinantly expressed and purified. Secondary-structure analysis using circular-dichroism spectroscopy indicated that MtbCarD contains ∼60% α-helix, ∼7% β-sheet and ∼33% random-coil structure. The C-terminal domain of MtbCarD (CarD83-161) was crystallized and its X-ray structure was determined at 2.1Å resolution. CarD83-161 forms a distorted Y-shaped structure containing bundles of three helices connected by a loop. The residues forming the distorted Y-shaped structure are highly conserved in CarD sequences from other mycobacterial species. Comparison of the CarD83-161 structure with the recently determined full-length M. tuberculosis and T. thermophilus CarD crystal structures revealed structural differences in residues 141-161 of the C-terminal domain of the CarD83-161 structure. The structural changes in the CarD83-161 structure occurred owing to proteolysis and crystallization artifacts.

Original languageEnglish (US)
Pages (from-to)160-165
Number of pages6
JournalActa Crystallographica Section F:Structural Biology Communications
Volume70
Issue number2
DOIs
StatePublished - Feb 2014

Keywords

  • CarD binding analysis
  • Mycobacterium tuberculosis CarD
  • Thermus aquaticus RNAP β subunit

ASJC Scopus subject areas

  • Biophysics
  • Structural Biology
  • Biochemistry
  • Genetics
  • Condensed Matter Physics

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