Trojan Horse Siderophore Conjugates Induce Pseudomonas aeruginosa Suicide and Qualify the TonB Protein as a Novel Antibiotic Target

Abstract

Rising infection rates with multidrug-resistant pathogens calls for antibiotics with novel modes of action. Herein, we identify the inner membrane protein TonB, a motor of active uptake in Gram-negative bacteria, as a novel target in antimicrobial therapy. The interaction of the TonB box of outer membrane transporters with TonB is crucial for the internalization of essential metabolites. We designed TonB box peptides and coupled them with synthetic siderophores in order to facilitate their uptake into bacteria in up to 32 synthetic steps. Three conjugates repressed the growth of Pseudomonas aeruginosa cells unable to produce their own siderophores, with minimal inhibitory concentrations between 0.1 and 0.5 μM. The transporters mediating uptake of these compounds were identified as PfeA and PirA. The study illustrates a variant of cellular suicide where a transporter imports its own inhibitor and demonstrates that artificial siderophores can import cargo with molecular weights up to 4 kDa.

Molecular Signatures of the Eagle Effect Induced by the Artificial Siderophore Conjugate LP-600 in E. coli

“…These data are also congruent with proteomic data from a study of LP-600 in P. aeruginosa. 60 Here, the import of the unconjugated MECAM core also induced the expression of the catechol-type outer membrane transporters PfeA and PirA, whereas LP-600 led to an increase in the expression of pfeA and ampC, the gene conferring ampicillin resistance. Interestingly, the expression of genes encoding the endogenous siderophores pyochelin and pyoverdine was repressed in the presence of the artificial siderophores in P. aeruginosa, whereas siderophore biosynthesis genes were strongly upregulated in this study.…”

Siderophore conjugates to combat antibiotic-resistant bacteria

“…Ultimately this work further demonstrated the ability of MECAM and DOTAM to transport a large cargo (kilodalton range) into bacterial cells. 111 Further, this is the first example of antimicrobial agents that disrupt a key periplasmic protein–protein interaction in a “cellular-suicide” manner. 111 This seminal work paves the way for the utilisation of siderophores in ‘out-of-the-box’ ways, as well as providing new antimicrobial targets which are urgently needed.…”

“…111 Further, this is the first example of antimicrobial agents that disrupt a key periplasmic protein–protein interaction in a “cellular-suicide” manner. 111 This seminal work paves the way for the utilisation of siderophores in ‘out-of-the-box’ ways, as well as providing new antimicrobial targets which are urgently needed. We await further work in this field, potentially investigating peptidomimetics or even small-molecular inhibitors of this key protein–protein interaction.…”

“…In innovative work, Brönstrup, Schalk and colleagues have demonstrated the first use of a TonB box peptide conjugated to a siderophore to induce TonB inhibition and subsequent bacterial suicide. 111 Their recent publication repurposed previously described synthetic siderophores MECAM and DOTAM (Fig. 8).…”

“…The TonB box is a component of TonB dependent transporters (TBDTs) that extends off the periplasmic domain of the TonB protein into the periplasm and is a sequence of five to seven amino acids which show some conservation. 111 This work investigated the potential for the disruption of the TBDT-TonB interaction as novel target in antimicrobial chemotherapy. 111 To execute this, a series of synthetic TonB box peptides were designed as competitive inhibitors of protein-protein interactions with several P. aeruginosa TBDTs (FpvA, HasR and PfeA) and were conjugated to the aforementioned triscatecholate synthetic siderophores.…”