Sujet de M2R 2016
Characterization of key enzymes involved in lipid processing in mycobacteria
Hédia Marrakchi [email protected] and Anne Lemassu [email protected]
Equipe Daffé, IPBS-CNRS-UPS Toulouse
Tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb) is a global pandemic, killing
someone approximately every 25 seconds and nearly 1.4 million people every year. The development
of new antibiotics effective against drug-resistant strains is one of the major priorities to fight against
TB, notably through the discovery of new pharmacological targets in the tubercle bacillus. This
approach requires to decipher key metabolic steps of the tubercle bacillus.
The cell envelope of Mtb is extremely rich in lipids that play a key role in its architecture and
permeability. The importance of lipids in mycobacteria is illustrated by the vast number of genes
dedicated to lipid metabolism, as revealed by analysis of the Mtb complete genome sequence. The
biogenesis of this envelope has been well studied in mycobacteria and is a validated target of several
antituberculous drugs. However, lipid outcome has not been investigated thoroughly and may have
specific relevance for Mtb physiology during survival in the host environment. Indeed, the products of
lipid metabolism may represent key compounds required for Mtb survival within the host tissues,
where the bacillus encounters various stress conditions (acidification, nitric oxide, hypoxia...).
The main objective of this project is to explore the mycobacterial lipid metabolic pathway(s)
by deciphering associated intermediates and identifying the implicated key enzymes that could be
developed as promising drug targets for antituberculous therapies. Mutants in the relevant gene
candidates will be constructed and their lipidomic profiles determined based on methods recently
developed in the lab. New analytical methods of the lipid intermediates will be developed, if needed,
for the study of the compounds that would accumulate in the mutants and when stress conditions are
applied to bacteria.
Techniques: mycobacterial cultures; search for candidate genes (bioinformatics) ; construction of
deletion mutants ; lipidomics (qualitative and quantitative analysis of extractable lipids by HPTLC, GCMS, UPLC-MS); lipid purification; structural analyses (controlled chemical degradations, NMR, MS).
References:
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Marrakchi H, Lanéelle MA, Daffé M. Mycolic acids: structures, biosynthesis,
and beyond. Chem Biol. 2014 Jan 16;21(1):67-85.