Molecular, kinetic, thermodynamic, and structural analyses of Mycobacterium tuberculosis hisD-encoded metal-dependent dimeric histidinol dehydrogenase (EC 1.1.1.23)

José E S Nunes, Rodrigo G. Ducati, Ardala Breda, Leonardo A. Rosado, Bibiana M. De Souza, Mario S. Palma, Diógenes S. Santos, Luiz A. Basso

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

The emergence of drug-resistant strains of Mycobacterium tuberculosis, the major causative agent of tuberculosis (TB), and the deadly HIV-TB co-infection have led to an urgent need for the development of new anti-TB drugs. The histidine biosynthetic pathway is present in bacteria, archaebacteria, lower eukaryotes and plants, but is absent in mammals. Disruption of the hisD gene has been shown to be essential for M. tuberculosis survival. Here we present cloning, expression and purification of recombinant hisD-encoded histidinol dehydrogenase (MtHisD). N-terminal amino acid sequencing and electrospray ionization mass spectrometry analyses confirmed the identity of homogeneous MtHisD. Analytical gel filtration, metal requirement analysis, steady-state kinetics and isothermal titration calorimetry data showed that homodimeric MtHisD is a metalloprotein that follows a Bi Uni Uni Bi Ping-Pong mechanism. pH-rate profiles and a three-dimensional model of MtHisD allowed proposal of amino acid residues involved in either catalysis or substrate(s) binding.

Original languageEnglish (US)
Pages (from-to)143-153
Number of pages11
JournalArchives of Biochemistry and Biophysics
Volume512
Issue number2
DOIs
StatePublished - Aug 15 2011
Externally publishedYes

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Thermodynamics
Mycobacterium tuberculosis
Tuberculosis
Metals
Metalloproteins
Amino Acids
Electrospray ionization
Kinetics
Mammals
Cloning
Calorimetry
Titration
Histidine
Pharmaceutical Preparations
Catalysis
Purification
Mass spectrometry
Bacteria
Electrospray Ionization Mass Spectrometry
Genes

Keywords

  • Enzyme mechanism
  • Histidinol dehydrogenase
  • Metalloenzyme
  • Molecular model
  • Mycobacterium tuberculosis
  • Thermodynamic binding parameters

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

Molecular, kinetic, thermodynamic, and structural analyses of Mycobacterium tuberculosis hisD-encoded metal-dependent dimeric histidinol dehydrogenase (EC 1.1.1.23). / Nunes, José E S; Ducati, Rodrigo G.; Breda, Ardala; Rosado, Leonardo A.; De Souza, Bibiana M.; Palma, Mario S.; Santos, Diógenes S.; Basso, Luiz A.

In: Archives of Biochemistry and Biophysics, Vol. 512, No. 2, 15.08.2011, p. 143-153.

Research output: Contribution to journalArticle

Nunes, José E S ; Ducati, Rodrigo G. ; Breda, Ardala ; Rosado, Leonardo A. ; De Souza, Bibiana M. ; Palma, Mario S. ; Santos, Diógenes S. ; Basso, Luiz A. / Molecular, kinetic, thermodynamic, and structural analyses of Mycobacterium tuberculosis hisD-encoded metal-dependent dimeric histidinol dehydrogenase (EC 1.1.1.23). In: Archives of Biochemistry and Biophysics. 2011 ; Vol. 512, No. 2. pp. 143-153.
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