Thiocillin and micrococcin exploit the ferrioxamine receptor of Pseudomonas aeruginosa for uptake

Abstract

Background Thiopeptides are a class of antibiotics that are active against Gram-positive bacteria and inhibit translation. They were considered inactive against Gram-negative bacteria due to their inability to cross the outer membrane. However, we discovered previously that a member of this class, thiostrepton (TS), has activity against Pseudomonas aeruginosa and Acinetobacter baumannii under iron-limiting conditions. TS hijacks the pyoverdine siderophore receptors of P. aeruginosa to cross the outer membrane and synergizes with iron chelators. Objectives To test other thiopeptides for antimicrobial activity against P. aeruginosa and determine their mechanism of uptake, action and spectrum of activity. Methods Eight thiopeptides were screened in chequerboard assays against a mutant of P. aeruginosa PA14 lacking both pyoverdine receptors. Thiopeptides that retain activity against a pyoverdine receptor-null mutant may use alternative siderophore receptors for entry. Susceptibility testing against siderophore receptor mutants was used to determine thiopeptide mechanism of uptake. Results The thiopeptides thiocillin (TC) and micrococcin (MC) use the ferrioxamine siderophore receptor (FoxA) for uptake and inhibit the growth of P. aeruginosa at low micromolar concentrations. The activity of TC required the TonB-ExbBD system used to energize siderophore uptake. TC acted through its canonical mechanism of action of translation inhibition. Conclusions Multiple thiopeptides have antimicrobial activity against P. aeruginosa, countering the historical assumption that they cannot cross the outer membrane. These results demonstrate the potential for thiopeptides to act as antipseudomonal antibiotics.

Essential role of membrane vesicles for biological activity of the bacteriocin micrococcin P1

“…In contrast, there was no antimicrobial activity in extracts from A. baumannii after association, indicating that MP1‐loaded MVs cannot deliver MP1 to that Gram‐negative bacterium, most likely due to the presence of the outer membrane. This is noteworthy, as pure MP1 also lacks activity toward Gram‐negative bacteria, which is believed to be due to the inability of MP1 to pass the outer membrane, except when specific receptors are present (Chan & Burrows, 2021 ).…”

Antimicrobial Bacillus: Metabolites and Their Mode of Action

“…Thiocillin (38) ( Figure 10 ), produced by B. cereus ATCC 14579, has been previously reported to only target gram-positive bacteria but has recently been shown to also target gram-negative bacteria [ 112 ]. Its mechanism on gram-positive bacteria works by targeting the 50S ribosome and inhibiting its role in protein synthesis [ 113 ].…”

Pseudomonas aeruginosa FpvB is a high-affinity transporter for xenosiderophores ferrichrome and ferrioxamine B

“…The copyright holder for this preprint this version posted September 20, 2022. ; https://doi.org/10.1101/2022.09.20.508722 doi: bioRxiv preprint Previously we showed that the large thiopeptide antibiotics thiostrepton (TS) and thiocillin (TC) use TBDTs to enter P. aeruginosa to access their cytoplasmic target, the ribosome. TS exploits the pyoverdine transporters, FpvA and FpvB, while thiocillin uses the ferrioxamine transporter, FoxA (Figure 1) (Chan et al, 2020;Chan and Burrows, 2021;Ranieri et al, 2019).…”

“…Together with the inner membrane proteins ExbB-ExbD, TonB harnesses the proton motive force to actively transport ligands through the transporters via a mechanism that remains incompletely understood (Klebba et al, 2021; Noinaj et al, 2010). Although TonB-dependent transporters (TBDTs) are considered ligand-specific, they can be exploited by antimicrobial compounds, bacteriophages, and bacteriocins for uptake, making them of interest for drug delivery across the outer membrane of Gram negatives (Chan and Burrows, 2021; Ferguson et al, 2000; Luna et al, 2020; Luscher et al, 2018; Mathavan et al, 2014; Rabsch et al, 2007; Ranieri et al, 2019; Salomon and Farias, 1993; White et al, 2017).…”

“…Previously we showed that the large thiopeptide antibiotics thiostrepton (TS) and thiocillin (TC) use TBDTs to enter P. aeruginosa to access their cytoplasmic target, the ribosome. TS exploits the pyoverdine transporters, FpvA and FpvB, while thiocillin uses the ferrioxamine transporter, FoxA (Figure 1) (Chan et al, 2020; Chan and Burrows, 2021; Ranieri et al, 2019). Here we further characterized the interaction of TS with the pyoverdine transporters.…”