title: Biochemical and physiological characterization of the GTP-binding protein Obg of Mycobacterium tuberculosis.
abstract: BACKGROUND
  Obg is a highly conserved GTP-binding protein that has homologues in bacteria, archaea and eukaryotes. In bacteria, Obg proteins are essential for growth, and they participate in spore formation, stress adaptation, ribosome assembly and chromosomal partitioning. This study was undertaken to investigate the biochemical and physiological characteristics of Obg in Mycobacterium tuberculosis, which causes tuberculosis in humans.
  
  
  RESULTS
  We overexpressed M. tuberculosis Obg in Escherichia coli and then purified the protein. This protein binds to, hydrolyzes and is phosphorylated with GTP. An anti-Obg antiserum, raised against the purified Obg, detects a 55 kDa protein in immunoblots of M. tuberculosis extracts. Immunoblotting also discloses that cultured M. tuberculosis cells contain increased amounts of Obg in the late log phase and in the stationary phase. Obg is also associated with ribosomes in M. tuberculosis, and it is distributed to all three ribosomal fractions (30 S, 50 S and 70 S). Finally, yeast two-hybrid analysis reveals that Obg interacts with the stress protein UsfX, indicating that M. tuberculosis Obg, like other bacterial Obgs, is a stress related protein.
  
  
  CONCLUSIONS
  Although its GTP-hydrolyzing and phosphorylating activities resemble those of other bacterial Obg homologues, M. tuberculosis Obg differs from them in these respects: (a) preferential association with the bacterial membrane; (b) association with all three ribosomal subunits, and (c) binding to the stress protein UsfX, rather than to RelA. Generation of mutant alleles of Obg of M. tuberculosis, and their characterization in vivo, may provide additional insights regarding its role in this important human pathogen.
authors.affiliation: Regional Academic Health Center and Department of Microbiology and Immunology, The University of Texas Health Science Center at San Antonio, Edinburg, Texas 78541, USA.
authors.forename: Smitha J
authors.initials: SJ
authors.lastname: Sasindran
authors.forename: Sankaralingam
authors.initials: S
authors.lastname: Saikolappan
authors.forename: Virginia L
authors.initials: VL
authors.lastname: Scofield
authors.forename: Subramanian
authors.initials: S
authors.lastname: Dhandayuthapani
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004269:DNA, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D012270:Ribosomes / Q000378:metabolism; D020798:Two-Hybrid System Techniques
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D008565:Membrane Proteins; C090542:Obg GTP-binding protein, Bacteria; C540214:UsfX protein, Mycobacterium tuberculosis; D019204:GTP-Binding Proteins