title: Deletion and overexpression studies on DacB2, a putative low molecular mass penicillin binding protein from Mycobacterium tuberculosis H(37)Rv.
abstract: Mycobacterium tuberculosis genome encodes several high and low molecular mass penicillin binding proteins. One such low molecular mass protein is DacB2 encoded by open reading frame Rv2911 of M. tuberculosis which is predicted to play a role in peptidoglycan synthesis. In this study we have tried to gain an insight into the role of this accessory cell division protein in mycobacterial physiology by performing overexpression and deletion studies. The overproduction of DacB2 in non-pathogenic, fast growing mycobacterium Mycobacterium smegmatis mc(2)155 resulted in reduced growth, an altered colony morphology, a defect in sliding motility and biofilm formation. A point mutant of DacB2 was made wherein the active site serine residue was mutated to cysteine to abolish the penicillin binding function of protein. The overexpression of mutant protein showed similar results indicating that the effects produced were independent of protein's penicillin binding function. The gene encoding DacB2 was deleted in M. tuberculosis by specialized transduction method. The deletion mutant showed reduced growth in Sauton's medium under acidic and low oxygen availability. The in vitro infection studies with THP-1 cells showed increased intracellular survival of dacB2 mutant compared to parent and complemented strains. The colony morphology and antibiotic sensitivity of mutant and wild-type strains were similar.
authors.affiliation: Department of Immunology, Tuberculosis Research Centre (Indian Council of Medical Research), Mayor V. R. Ramanathan Road, Chetpet, Chennai 600031, India.
authors.forename: Neema
authors.initials: N
authors.lastname: Bourai
authors.forename: William R
authors.initials: WR
authors.lastname: Jacobs
authors.forename: Sujatha
authors.initials: S
authors.lastname: Narayanan
mesh_terms: D000995:Antitubercular Agents / Q000494:pharmacology; D002460:Cell Line; D003470:Culture Media / Q000737:chemistry; D017353:Gene Deletion; D015870:Gene Expression; D005816:Genetic Complementation Test; D006801:Humans; D008124:Locomotion; D008826:Microbial Sensitivity Tests; D050296:Microbial Viability; D009000:Monocytes / Q000382:microbiology; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D020125:Mutation, Missense; D020102:Mycobacterium smegmatis / Q000235:genetics / Q000254:growth & development / Q000502:physiology; D009169:Mycobacterium tuberculosis / Q000235:genetics* / Q000254:growth & development / Q000502:physiology*; D046915:Penicillin-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D010406:Penicillins / Q000378:metabolism
chemical_list: D000995:Antitubercular Agents; D003470:Culture Media; D050505:Mutant Proteins; D046915:Penicillin-Binding Proteins; D010406:Penicillins