Phosphorylation of the Mycobacterium tuberculosis β-Ketoacyl-Acyl Carrier Protein Reductase MabA Regulates Mycolic Acid Biosynthesis
Open accessAbstract
Mycolic acids are key cell wall components for the survival, pathogenicity, and antibiotic resistance of the human tubercle bacillus. Although it was thought that Mycobacterium tuberculosis tightly regulates their production to adapt to prevailing environmental conditions, the molecular mechanisms governing mycolic acid biosynthesis remained extremely obscure. Meromycolic acids, the direct precursors of mycolic acids, are synthesized by a type II fatty acid synthase from acyl carrier protein-bound substrates that are extended iteratively, with a reductive cycle in each round of extension, the second step of which is catalyzed by the essential beta-ketoacyl-acyl carrier protein reductase, MabA. In this study, we investigated whether post-translational modifications of MabA might represent a strategy employed by M. tuberculosis to regulate mycolic acid biosynthesis. Indeed, we show here that MabA was efficiently phosphorylated in vitro by several M. tuberculosis Ser/Thr protein kinases, including PknB, as well as in vivo in mycobacteria. Mass spectrometric analyses using LC-ESI/MS/MS and site-directed mutagenesis identified three phosphothreonines, with Thr(191) being the primary phosphor-acceptor. A MabA_T191D mutant, designed to mimic constitutive phosphorylation, exhibited markedly decreased ketoacyl reductase activity compared with the wild-type protein, as well as impaired binding of the NADPH cofactor, as demonstrated by fluorescence spectroscopy. The hypothesis that phosphorylation of Thr(191) alters the enzymatic activity of MabA, and subsequently mycolic acid biosynthesis, was further supported by the fact that constitutive overexpression of the mabA_T191D allele in Mycobacterium bovis BCG strongly impaired mycobacterial growth. Importantly, conditional expression of the phosphomimetic MabA_T191D led to a significant inhibition of de novo biosynthesis of mycolic acids. This study provides the first information on the molecular mechanism(s) involved in mycolic acid regulation through Ser/Thr protein kinase-dependent phosphorylation of a type II fatty acid synthase enzyme.
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The FAS-II enzymes are all inhibited through phosphorylation by PknA, PknB and other STPKs.
“The FAS-II enzymes are all inhibited through phosphorylation by PknA, PknB and other STPKs89–93.”
From The mycobacterial cell envelope — a moving target (2020) · cites this paper as [93]
The MabA phosphorylation results and Discussion show that several FAS-II enzymes are STPK substrates; MabA is phosphorylated in vitro by PknA, PknB, and other kinases, and a phosphomimetic MabA variant has reduced activity. But the Discussion says the effect of phosphorylation on mtFabD activity is unknown and whether HadABC and InhA are STPK substrates remains unresolved. The paper therefore does not support the claim that all FAS-II enzymes are inhibited through phosphorylation. (LLM, conf=0.90)
Experimental — accuracy not yet measuredPknH, PknD, PknL and PknF can also phosphorylate various FAS-II enzymes.
“PknH, PknD, PknL and PknF can also phosphorylate various FAS-II enzymes.”
From The mycobacterial cell envelope — a moving target (2020) · cites this paper as [93]
The section “MabA Is Phosphorylated in Vitro by Multiple Ser/Thr Kinases” reports that PknD and PknL phosphorylated MabA, a FAS-II enzyme. It reports no phosphorylation of MabA by PknH or PknF. The Discussion describes phosphorylation of other FAS-II enzymes but does not establish that PknH or PknF phosphorylate them, so the claim about all four named kinases is not supported. (LLM, conf=0.86)
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Cites
Critical Residues for Structure and Catalysis in Short-chain Dehydrogenases/Reductases
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Research works printed but not linked: 3
- Kremer, L., Baulard, A. R., and Besra, G. S. (2000) in Molecular Genetics of Mycobacteria (Hatfull, G. F., and Jacobs, W. R., Jr., eds) pp. 173-190, ASM Press, Washington, DC
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The mycobacterial cell envelope — a moving target
Dulberger, C. L., Rubin, E. J., & Boutte, C. C. (2020) · Nature Reviews Microbiology · doi:10.1038/s41579-019-0273-7 · Cited by 365 · see citing works
- Authors
- Veyron-Churlet, R., Zanella-Cléon, I., Cohen-Gonsaud, M., Molle, V., & Kremer, L.
- Affiliations
- Lyon 1 Université (FR)
- Centre National de la Recherche Scientifique (FR)
- Corresponding author
- Molle, Virginie
- Kremer, Laurent
- Venue
- Journal of Biological Chemistry
- Publisher
- Elsevier BV
- Published
- 2010-04
- Volume
- 285
- Issue
- 17
- Pages
- 12714-12725
- ISSN
- 0021-9258 (ISSN-L)
- PMC
- PMC2857119
- OpenAlex
- W2012027216
- Cited by
- 80
- References
- 4
- Citations by year
- 2026: 2 · 2025: 4 · 2023: 1 · 2022: 2 · 2021: 3 · 2020: 6 · 2019: 4 · 2018: 4 · 2017: 8 · 2016: 5 · 2015: 5 · 2014: 9 · 2013: 5 · 2012: 8
- FWCI
- 5.60
- Type
- journal article
- Language
- en
- Open access
- hybrid
- doi.org
- Licence
- Locations
- Journal of Biological Chemistry (publishedVersion, cc-by)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- HAL (submittedVersion, cc-by)
- Topics
- Health Sciences › Medicine › Infectious Diseases › Tuberculosis Research and Epidemiology
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Biochemistry › Lipid metabolism and biosynthesis
- Health Sciences › Medicine › Pharmacology › Microbial Natural Products and Biosynthesis
- Keywords
- Mycobacterium tuberculosis, MabA, mycolic acid biosynthesis, type II fatty acid synthase, protein phosphorylation, serine/threonine kinase, PknB, phosphomimetic mutant, mycobacterial cell wall, Mycobacterium bovis BCG
- MeSH terms
- Alcohol Oxidoreductases, Humans, Mycobacterium bovis, Mycobacterium tuberculosis, Mycolic Acids, Oxidoreductases, Phosphorylation, Protein Processing, Post-Translational, Spectrometry, Fluorescence, Gene Expression Regulation, Bacterial, Gene Expression Regulation, Enzymologic, Mutagenesis, Site-Directed, Protein Serine-Threonine Kinases, 3-Oxoacyl-(Acyl-Carrier-Protein) Reductase
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