Dataset Information

Phosphoproteomics to uncover phosphorylation substrates changes when serine/threonine protein kinase PknA and PknB depletion

ABSTRACT: The Mycobacterium tuberculosis eukaryotic-like serine/threonine protein kinase (STPKs) PknA and PknB regulate several processes required for cell growth, both kinase have been found to be and can be investigated by using the conditional mutant. In the present study, we created the M. tuberculosis kinase PknA/ PknB depletion mutant and performed the phosphoproteomics to uncover phosphorylation substrates changes when PknA/ PknB depletion.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Mycobacterium Tuberculosis C Stg4006

TISSUE(S): Cell Culture

SUBMITTER: Saima Ahmed

LAB HEAD: Hanno Steen

PROVIDER: PXD015197 | Pride | 2020-05-27

REPOSITORIES: Pride

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Publications

Protein kinases PknA and PknB independently and coordinately regulate essential Mycobacterium tuberculosis physiologies and antimicrobial susceptibility.

Zeng Jumei J Platig John J Cheng Tan-Yun TY Ahmed Saima S Skaf Yara Y Potluri Lakshmi-Prasad LP Schwartz Daniel D Steen Hanno H Moody D Branch DB Husson Robert N RN

PLoS pathogens 20200407 4

The Mycobacterium tuberculosis Ser/Thr protein kinases PknA and PknB are essential for growth and have been proposed as possible drug targets. We used a titratable conditional depletion system to investigate the functions of these kinases. Depletion of PknA or PknB or both kinases resulted in growth arrest, shortening of cells, and time-dependent loss of acid-fast staining with a concomitant decrease in mycolate synthesis and accumulation of trehalose monomycolate. Depletion of PknA and/or PknB ...[more]

PMID: 32255801

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Project description:Different transcriptome, metabolome, and phosphoproteome studies have already indicated a regulatory role of Ser/Thr phosphorylation in the central metabolism of S. aureus. However, it remains still unknown how this exactly tunes its metabolism. For that reason, Extreme Pathway Analysis was used to understand metabolic flux regulation by Ser/Thr phosphorylation. YANAsquare software and YANAvergence routine were chosen to calculate pathway activities of WT, ΔpknB, Δstp, and double mutant S. aureus NewmanHG strains. Gene expression data of S. aureus NewmanHG were collected taking the wild type and mutations of kinase (pknB), phosphatase (stp) or both (pknB/stp) phenotypes. The resulting flux changes were modelled with YANAvergence taking the different gene expression data and an established genome scale model of S. aureus metabolic modes as basis. The fluxes of the different strains were next calculated, taking the gene expression data into account. A concerted metabolic change of fluxes was observed. In particular the pathways for peptidoglycan, nucleotide, and aromatic amino acid synthesis as well as catabolism involving aspartate transaminase (GOT) changed significantly compared to the wild type. Single mutations of pknB and stp were compared to illustrate their different contribution to the phenotype, e.g. pyrimidine synthesis is dramatically impaired by pknB deletion but much less by loss of stp. In the double knock out strain, there is higher activity of peptidoglycan, purine, and aromatic amino acids synthesis from glucose, but lower activity of pyrimidine synthesis from glucose compared to WT. The results suggest a probable regulatory role of Ser/Thr phosphorylation in amino acid catabolism and the glycolysis/gluconeogenesis switch to provide the cell with sufficient cell wall components, nucleotides, and aromatic amino acids. Yvck is suggested to serve as regulatory protein linking Ser/Thr phosphorylation to the previous results of S. aureus metabolism. Moreover, Yvck seems in particular to be involved in the switch between glycolysis and gluconeogenesis. However, further studies based on experimental approaches are needed to demonstrate that Ser/Thr phosphorylation and Yvck are necessary for a correct cell growth under gluconeogenic conditions. The metabolic model allows not only a comprehensive representation of these changes for all involved pathways in the four strains, it further points out the function of the previously unannotated ycvk operon.. The relevance of these specific metabolic pathway adaptations regarding the pathogenicity of S.aureus and their contributions to the infection process will be analyzed and discussed.

Dataset Information

Phosphoproteomics to uncover phosphorylation substrates changes when serine/threonine protein kinase PknA and PknB depletion

Publications

Protein kinases PknA and PknB independently and coordinately regulate essential Mycobacterium tuberculosis physiologies and antimicrobial susceptibility.

Similar Datasets

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