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ESTABLISHMENT: University of Lille

Laboratory of affiliation: UMRt 1158 BioEcoAgro

Scientific field, Speciality: DS10 | Food Biotechnology, Food Science, Physiology

Thesis director: LECLERE Valérie, Pr, valerie.leclere@univ-lille.fr

Co-supervisor:  DUBAN Matthieu, IgR, matthieu.duban@univ-lille.fr

Affiliate programme(s): ex. labex, ERC, Horizon Europe, etc

Planned (co)-funding (mention: in progress/obtained): Région HdF, ED SMRE

Title of the thesis:

Chassiclip: Creation of bacterial chassis strains for the heterologous production of lipopeptides and optimization of production, extraction and purification processes

THESIS SUBJECT:

Cyclic lipopeptides or CLiPs (Cyclic LipoPeptides) are microbial secondary metabolites with surfactant and antifungal properties that make it possible to consider their use in biocontrol applications. They can act through direct antagonism mechanisms towards the pathogen (biocides) or indirectly by inducing the host plant's systemic defence response (SDP) (Ongena and Jacques, 2008).

CLiPs are synthesised by different bacterial genera (notably Bacillus, Pseudomonas or Burkholderia) through non-ribosomal synthesis pathways, thanks to modular mega-enzymes called non-ribosomal peptide synthetases or NRPSs (Marahiel, 2016). NRPSs work as production chains to catalyse, step by step, the assembly of amino acids leading to the construction of non-ribosomal peptides. NRPS are encoded by very large genes, gathered in biosynthetic gene clusters (BGCs) of several tens of thousands of bases, which also contain non-NRPS genes essential for the synthesis of non-ribosomal peptides. Bioinformatics tools are now available to detect the presence of such BGCs in genomes and to predict the synthesis of CLiPs in silico (Pupin et al., 2018).

Several bottlenecks to the use of CLiPs in biocontrol have been identified. CLiPs are generally produced naturally in small quantities (at most a few tens of mg per litre of culture), and a single strain is often capable of co-producing several lipopeptides in a mixture (D'aes et al., 2014), which poses difficulties in terms of their extraction/purification. On the other hand, some BGCs are even cryptic, i.e. not expressed under laboratory culture conditions. One way of getting around these difficulties is therefore to use heterologous production, which consists of using a host strain into which BGCs containing biosynthesis genes are introduced. The proposed subject involves the molecular construction of chassis strains by genetic engineering, capable of receiving lipopeptide BGCs of various origins. Optimisation of the production, extraction and purification processes is envisaged from one or two models in order to obtain sufficient quantities to analyse their structure, understand their mode of action, and evaluate their activities at different scales (laboratory, greenhouse, field).

Expected date of recruitment: 1st October 2023

Contact:valerie.leclere@univ-lille.fr

Expected skills: microbiology, molecular biology, fermentation