title: Phosphorylation of pyruvate kinase A by protein kinase J leads to the altered growth and differential rate of intracellular survival of mycobacteria.
abstract: PknJ (Rv2088) is a serine/threonine protein kinase of mycobacteria which is present in Mycobacterium tuberculosis (MTB), but its gene is absent in Mycobacterium smegmatis (MS); a fast grower and nonpathogenic species of mycobacteria. The heterologous expression of MTB-specific PknJ in MS altered the growth of recombinant mycobacteria highlighting one of the characteristics of this protein. This nature of the protein was further confirmed when Mycobacterium bovis BCG (BCG) containing antisense copy of pknJ resulted in the increased growth of BCG. The real-time RNA quantification analysis pointed out toward increased expression of this protein during infection of THP-1 macrophage cells which further emphasized that the protein is essential for the intracellular survival of mycobacteria. The differential in gel electrophoresis (DIGE) data followed by mass spectroscopy suggested that PknJ is involved in regulation of pyruvate kinase A (Rv1617). Since pyruvate kinase (PK) A is one of the key enzymes which controls glycolytic cycle in mycobacteria, we looked for its interaction with PknJ during extracellular and intracellular growth of mycobacteria. In order to identify the specific residue(s) involved in post-translational modification, the phospho-null mutants of PK were generated, and their substrate specificities in response to PknJ were assessed through kinase assay. The findings thus underlined that the PK activity is predominantly dependent on the threonine residue at the 94(th) position and further suggested that this site may be plausible in intracellular survival of mycobacteria upon phosphorylation with PknJ.
authors.affiliation: Division of Microbiology and Academy of Scientific and Innovative Research, CSIR, Central Drug Research Institute, Lucknow, 226031, India.
authors.forename: Diwakar K
authors.initials: DK
authors.lastname: Singh
authors.forename: Pramod K
authors.initials: PK
authors.lastname: Singh
authors.forename: Sameer
authors.initials: S
authors.lastname: Tiwari
authors.forename: Susmita K
authors.initials: SK
authors.lastname: Singh
authors.forename: Ruma
authors.initials: R
authors.lastname: Kumari
authors.forename: Dinesh K
authors.initials: DK
authors.lastname: Tripathi
authors.forename: Kishore K
authors.initials: KK
authors.lastname: Srivastava
mesh_terms: D002460:Cell Line; D020869:Gene Expression Profiling; D006801:Humans; D050296:Microbial Viability; D009000:Monocytes / Q000382:microbiology; D009163:Mycobacterium bovis / Q000201:enzymology / Q000254:growth & development / Q000378:metabolism / Q000502:physiology; D020102:Mycobacterium smegmatis / Q000201:enzymology / Q000254:growth & development / Q000378:metabolism / Q000502:physiology; D009169:Mycobacterium tuberculosis / Q000201:enzymology* / Q000254:growth & development / Q000378:metabolism / Q000502:physiology*; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D011770:Pyruvate Kinase / Q000378:metabolism*
chemical_list: D011494:Protein Kinases; C106540:protein kinase J; D011770:Pyruvate Kinase