Itaconate utilisation by the human pathogen Pseudomonas aeruginosa requires uptake via the IctPQM TRAP transporter

Abstract Pseudomonas aeruginosa PA01 is one of the major causes of disease persistence and mortality in patients with lung pathologies, relying on various host metabolites as carbon and energy sources for growth. The ict-ich-ccl operon (pa0878, pa0882, pa0883) in PAO1 is required for growth on the host molecule itaconate, a C5-dicarboxylate. However, it is not known how itaconate is taken up into P. aeruginosa. Here we demonstrate that a genetically linked tripartite ATP-independent periplasmic (TRAP) transporter (pa0884-pa0886), which is homologous to the known C4-dicarboxylate binding TRAP system, is essential for growth on itaconate, but not for the closely related C4-dicarboxylate succinate. Using tryptophan fluorescence spectroscopy we demonstrate that the substrate binding protein, IctP (PA0884), binds itaconate, but still retains higher affinity for the related C4-dicarboxylates. The structures of IctP bound to itaconate (1.80 Å) and succinate (1.75 Å), revealed an enclosed ligand binding pocket with ion pairing interactions with the ligand carboxylates. The C2 methylene group that is the distinguishing feature of itaconate compared to succinate is accommodated by a unique change in the IctP binding site from a Leu to Val, which distinguishes it from closely related C4-dicarboxylate binding SBPs. Together these data suggest that this transporter, which we name IctPQM, has duplicated from a canonical C4-dicarboxylate transporter and its evolution towards itaconate specificity enables this pathogen to now access a key metabolite for persistence in the host. Competing Interest Statement The authors have declared no competing interest. Footnotes ↵* Joint first authors