title: MRA_1571 is required for isoleucine biosynthesis and improves Mycobacterium tuberculosis H37Ra survival under stress.
abstract: Threonine dehydratase is a pyridoxal 5-phosphate dependent enzyme required for isoleucine biosynthesis. Threonine dehydratase (IlvA) participates in conversion of threonine to 2-oxobutanoate and ammonia is released as a by-product. MRA_1571 is annotated to be coding for IlvA in Mycobacterium tuberculosis H37Ra (Mtb-Ra). We developed a recombinant (KD) Mtb-Ra strain by down-regulating IlvA. The growth studies on different carbon sources suggested reduced growth of KD compared to wild-type (WT), also, isoleucine concentration dependent KD growth restoration was observed. The expression profiling of IlvA suggested increased expression of IlvA during oxygen, acid and oxidative stress. In addition, KD showed reduced survival under pH, starvation, nitric oxide and peroxide stresses. KD was more susceptible to antimycobacterial agents such as streptomycin (STR), rifampicin (RIF) and levofloxacin (LVF), while, no such effect was noticeable when exposed to isoniazid. Also, an increase in expression of IlvA was observed when exposed to STR, RIF and LVF. The dye accumulation studies suggested increased permeability of KD to ethidium bromide and Nile Red as compared to WT. TLC and Mass studies confirmed altered lipid profile of KD. In summary down-regulation of IlvA affects Mtb growth, increases its susceptibility to stress and leads to altered cell wall lipid profile.
authors.affiliation: Microbiology Division, CSIR-Central Drug Research Institute, B.S. 10/1, Sector 10, Jankipuram Extension, Sitapur Road, Lucknow-226031, India.
authors.forename: Rishabh
authors.initials: R
authors.lastname: Sharma
authors.affiliation: Microbiology Division, CSIR-Central Drug Research Institute, B.S. 10/1, Sector 10, Jankipuram Extension, Sitapur Road, Lucknow-226031, India.
authors.forename: Deepa
authors.initials: D
authors.lastname: Keshari
authors.affiliation: Microbiology Division, CSIR-Central Drug Research Institute, B.S. 10/1, Sector 10, Jankipuram Extension, Sitapur Road, Lucknow-226031, India.
authors.forename: Kumar Sachin
authors.initials: KS
authors.lastname: Singh
authors.affiliation: Microbiology Division, CSIR-Central Drug Research Institute, B.S. 10/1, Sector 10, Jankipuram Extension, Sitapur Road, Lucknow-226031, India.
authors.forename: Shailendra
authors.initials: S
authors.lastname: Yadav
authors.affiliation: Microbiology Division, CSIR-Central Drug Research Institute, B.S. 10/1, Sector 10, Jankipuram Extension, Sitapur Road, Lucknow-226031, India.|Academy of Scientific and Industrial Research (AcSIR), New Delhi, India.
authors.forename: Sudheer Kumar
authors.initials: SK
authors.lastname: Singh
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002473:Cell Wall / Q000235:genetics / Q000378:metabolism; D007532:Isoleucine / Q000096:biosynthesis / Q000235:genetics; D050356:Lipid Metabolism / Q000502:physiology; D050296:Microbial Viability; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000378:metabolism; D018384:Oxidative Stress / Q000502:physiology*; D013913:Threonine Dehydratase / Q000235:genetics / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D007532:Isoleucine; D013913:Threonine Dehydratase