Structural basis for antibacterial peptide self-immunity by the bacterial ABC transporter McjD.

Weixiao Yuan Wahlgren,Armin Wagner,Kiran Bountra,Valentina Corradi,D Peter Tieleman,Sylvie Rebuffat,Kamel El Omari,Hassanul G Choudhury,Séverine Zirah,Gregor Hagelueken,Konstantinos Beis,Indran Mathavan,Olav Schiemann

doi:10.15252/embj.201797278

Abstract

Certain pathogenic bacteria produce and release toxic peptides to ensure either nutrient availability or evasion from the immune system. These peptides are also toxic to the producing bacteria that utilize dedicated ABC transporters to provide self‐immunity. The ABC transporter McjD exports the antibacterial peptide MccJ25 in Escherichia coli. Our previously determined McjD structure provided some mechanistic insights into antibacterial peptide efflux. In this study, we have determined its structure in a novel conformation, apo inward‐occluded and a new nucleotide‐bound state, high‐energy outward‐occluded intermediate state, with a defined ligand binding cavity. Predictive cysteine cross‐linking in E. coli membranes and PELDOR measurements along the transport cycle indicate that McjD does not undergo major conformational changes as previously proposed for multi‐drug ABC exporters. Combined with transport assays and molecular dynamics simulations, we propose a novel mechanism for toxic peptide ABC exporters that only requires the transient opening of the cavity for release of the peptide. We propose that shielding of the cavity ensures that the transporter is available to export the newly synthesized peptides, preventing toxic‐level build‐up.

Highlights

Certain pathogenic bacteria produce and release toxic peptides to ensure either nutrient availability or evasion from the immune system
In order to gain a detailed understanding of the conformations that McjD adopts during the transport cycle, we have determined its structure in the presence of the transition state analogue ADP-VO4 and in the absence of nucleotides, that is in the apo form, at 3.4 Aand 4.7 Aresolution, respectively (Fig 1B and C)
A 2017 The Authors nucleotide-bound state, high-energy intermediate outward-occluded. These findings highlight novel mechanistic details on how McjD provides cells with self-immunity against antibacterial peptides. Since these new conformations have neither been previously reported nor observed for ATP-binding cassette (ABC) transporters, we performed predictive cysteine cross-linking in E. coli inside-out vesicles (ISOVs) to verify their existence in the lipid environment

Summary

Introduction

Certain pathogenic bacteria produce and release toxic peptides to ensure either nutrient availability or evasion from the immune system. These peptides are toxic to the producing bacteria that utilize dedicated ABC transporters to provide self-immunity. Our previously determined McjD structure provided some mechanistic insights into antibacterial peptide efflux. Predictive cysteine cross-linking in E. coli membranes and PELDOR measurements along the transport cycle indicate that McjD does not undergo major conformational changes as previously proposed for multi-drug ABC exporters. Combined with transport assays and molecular dynamics simulations, we propose a novel mechanism for toxic peptide ABC exporters that only requires the transient opening of the cavity for release of the peptide. We propose that shielding of the cavity ensures that the transporter is available to export the newly synthesized peptides, preventing toxic-level build-up

Objectives

Methods

Results

Conclusion