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Semi-rational evolution of the Pseudomonas aeruginosa Rhamnosyltransferase 1 subunit A (RhlA) for the synthesis of industrially relevant rhamnolipids

Dulcey, Carlos Eduardo; de los Santos, Yossef Lopez; Déziel, Éric; Doucet, Nicolas (2017). Semi-rational evolution of the Pseudomonas aeruginosa Rhamnosyltransferase 1 subunit A (RhlA) for the synthesis of industrially relevant rhamnolipids In: 31st Annual Symposium of the Protein-Society, July 24-27, 2017, Montréal (Québec) Canada.

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Résumé

Rhamnolipids (RLs) are glycolipidic compounds produced by a few of bacterial species, especially Pseudomonas and Burkholderia spp. These compounds display excellent surfactant properties and environmental advantages. Nevertheless, their high production cost hampers their practical use in industry. In addition, bacterial strains that produce RLs generate a mixture of congeners with varying lipophilic chain lengths, therefore affecting their macromolecular behavior. Since the physicochemical characteristics of RLs are directly influenced by their molecular structure, modification or improvement of their surfactant properties can be achieved by controlling the length of their alkyl chains. RhlA acts as a key enzyme in the RL biosynthesis pathway. The enzyme catalyzes the esterification reaction between two units of hydroxylated fatty acids to form a dimer, ß-3-(3-hydroxyalkanoyl) alkanoic acid (HAA), the dilipid precursor of RLs. HAA biosynthesis is the rate-limiting step in RL biosynthesis. Here, we present a semi-rational evolution approach to engineer RhlA from P. aeruginosa to increase in vivo HAA production and to modulate substrate selectivity. Using a structural homology model of the enzyme, we predicted a number of substrate-interacting residues and performed intragenic suppression-type mutagenesis to increase the catalytic efficiency of RhlA. We also employed a chimeric approach to identify protein domains involved in enzyme selectivity, in addition to performing site-directed mutagenesis on residues located in the substrate binding pocket to modulate RL congener biosynthesis. Our results provide evidence that protein engineering approaches can be efficiently employed to improve RL production in P. aeruginosa.

Type de document: Document issu d'une conférence ou d'un atelier
Informations complémentaires: Affiche scientifique POS330 Protein Science 26 (suppl. 1): 68
Mots-clés libres: -
Centre: Centre INRS-Institut Armand Frappier
Date de dépôt: 22 mars 2019 15:33
Dernière modification: 22 mars 2019 15:33
URI: http://espace.inrs.ca/id/eprint/6721

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