Uptake Mechanisms and Regulatory Responses to MECAM- and DOTAM-Based Artificial Siderophores and Their Antibiotic Conjugates in Pseudomonas aeruginosa

authored by

Sarah Fritsch, Véronique Gasser, Carsten Peukert, Lukas Pinkert, Lauriane Kuhn, Quentin Perraud, Vincent Normant, Mark Brönstrup, Isabelle J. Schalk

Abstract

The development of new antibiotics against Gram-negative bacteria has to deal with the low permeability of the outer membrane. This obstacle can be overcome by utilizing siderophore-dependent iron uptake pathways as entrance routes for antibiotic uptake. Iron-chelating siderophores are actively imported by bacteria, and their conjugation to antibiotics allows smuggling the latter into bacterial cells. Synthetic siderophore mimetics based on MECAM (1,3,5-N,N′,N″-tris-(2,3-dihydroxybenzoyl)-triaminomethylbenzene) and DOTAM (1,4,7,10-tetrakis(carbamoylmethyl)-1,4,7,10-tetraazacyclododecane) cores, both chelating iron via catechol groups, have been recently applied as versatile carriers of functional cargo. In the present study, we show that MECAM and the MECAM-ampicillin conjugate 3 transport iron into Pseudomonas aeruginosa cells via the catechol-type outer membrane transporters PfeA and PirA and DOTAM solely via PirA. Differential proteomics and quantitative real-time polymerase chain reaction (qRT-PCR) showed that MECAM import induced the expression of pfeA, whereas 3 led to an increase in the expression of pfeA and ampc, a gene conferring ampicillin resistance. The presence of DOTAM did not induce the expression of pirA but upregulated the expression of two zinc transporters (cntO and PA0781), pointing out that bacteria become zinc starved in the presence of this compound. Iron uptake experiments with radioactive ⁵⁵Fe demonstrated that import of this nutrient by MECAM and DOTAM was as efficient as with the natural siderophore enterobactin. The study provides a functional validation for DOTAM- and MECAM-based artificial siderophore mimetics as vehicles for the delivery of cargo into Gram-negative bacteria.

Organisation(s)

Centre of Biomolecular Drug Research (BMWZ)

External Organisation(s)

University of Strasbourg
Helmholtz Centre for Infection Research (HZI)
Centre national de la recherche scientifique (CNRS)

Type

Article

Journal

ACS infectious diseases

Volume

8

Pages

1134-1146

No. of pages

13

ISSN

2373-8227

Publication date

10.06.2022

Publication status

Published

Peer reviewed

Yes

ASJC Scopus subject areas

Infectious Diseases

Sustainable Development Goals

SDG 3 - Good Health and Well-being

Electronic version(s)

https://doi.org/10.1021/acsinfecdis.2c00049 (Access: Closed)