The Mycobacterium tuberculosis protein kinase K modulates activation of transcription from the promoter of mycobacterial monooxygenase operon through phosphorylation of the transcriptional regulator VirS.
Mycobacterium tuberculosis encodes for 11 eukaryotic-like serine/threonine protein kinases. Genetic and biochemical studies show that these kinases regulate various cellular processes including cell shape and morphology, glucose and glutamine transport, phagosome-lysosome fusion and the expression, and/or activity of transcription factors. PknK is the largest predicted serine/threonine protein kinase in M. tuberculosis. Here, we have cloned, overexpressed, and purified protein kinase K (PknK) to near homogeneity and show that its ability to phosphorylate proteins is dependent on the invariant lysine (Lys(55)), and on two conserved threonine residues present in its activation loop. Despite being devoid of any apparent transmembrane domain, PknK is localized to the cell wall fraction, suggesting probable anchoring of the kinase to the cell membrane region. The pknK gene is located in the vicinity of the virS gene, which is known to regulate the expression of the mycobacterial monooxygenase (mymA) operon. We report here for the first time that VirS is in fact a substrate of PknK. In addition, four of the proteins encoded by mymA operon are also found to be substrates of PknK. Results show that VirS is a bona fide substrate of PknK in vivo, and PknK-mediated phosphorylation of VirS increases its affinity for mym promoter DNA. Reporter assays reveal that PknK modulates VirS-mediated stimulation of transcription from the mym promoter. These findings suggest that the expression of mymA operon genes is regulated through PknK-mediated phosphorylation of VirS.
Kumar
Pawan
P
National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110 067, India.
Kumar
Devanand
D
Parikh
Amit
A
Rananaware
Dimple
D
Gupta
Meetu
M
Singh
Yogendra
Y
Nandicoori
Vinay Kumar
VK
D000595:Amino Acid Sequence; D002473:Cell Wall / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D006899:Mixed Function Oxygenases / Q000235:genetics / Q000378:metabolism; D008954:Models, Biological; D008969:Molecular Sequence Data; D009169:Mycobacterium tuberculosis / Q000201:enzymology*; D009876:Operon; D010587:Phagocytosis; D010750:Phosphoproteins / Q000096:biosynthesis / Q000502:physiology*; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000502:physiology*; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic
D010750:Phosphoproteins; D004247:DNA; D006899:Mixed Function Oxygenases; D017346:Protein Serine-Threonine Kinases