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Structure of an antibacterial peptide ATP-binding cassette transporter in a novel outward occluded state
DOI:
10.1073/pnas.1320506111
PMID:
24920594
Authors:
Hassan
Choudhury
(Imperial College London)
,
Zhen
Tong
(University of Cambridge)
,
Indran
Mathavan
(Imperial College London)
,
Yanyan
Li
(Centre National de la Recherche Scientifique)
,
So
Iwata
(Diamond Light Source)
,
Séverine
Zirah
(Centre National de la Recherche Scientifique)
,
Sylvie
Rebuffat
(Centre National de la Recherche Scientifique)
,
Hendrik W.
Van Veen
(University of Cambridge)
,
Konstantinos
Beis
(Imperial College London; Diamond Light Source; Research Complex at Harwell)
Co-authored by industrial partner:
No
Type:
Journal Paper
Journal:
Proceedings Of The National Academy Of Sciences
, VOL 111 (25)
, PAGES 9145 - 9150
State:
Published (Approved)
Published:
June 2014
Diamond Proposal Number(s):
6384

Abstract: Enterobacteriaceae produce antimicrobial peptides for survival under nutrient starvation. Microcin J25 (MccJ25) is an antimicrobial peptide with a unique lasso topology. It is secreted by the ATP-binding cassette (ABC) exporter McjD, which ensures self-immunity of the producing strain through efficient export of the toxic mature peptide from the cell. Here we have determined the crystal structure of McjD from Escherichia coli at 2.7-Å resolution, which is to the authors knowledge the first structure of an antibacterial peptide ABC transporter. Our functional and biochemical analyses demonstrate McjD-dependent immunity to MccJ25 through efflux of the peptide. McjD can directly bind MccJ25 and displays a basal ATPase activity that is stimulated by MccJ25 in both detergent solution and proteoliposomes. McjD adopts a new conformation, termed nucleotide-bound outward occluded. The new conformation defines a clear cavity; mutagenesis and ligand binding studies of the cavity have identified Phe86, Asn134, and Asn302 as important for recognition of MccJ25. Comparisons with the inward-open MsbA and outward-open Sav1866 structures show that McjD has structural similarities with both states without the intertwining of transmembrane (TM) helices. The occluded state is formed by rotation of TMs 1 and 2 toward the equivalent TMs of the opposite monomer, unlike Sav1866 where they intertwine with TMs 36 of the opposite monomer. Cysteine cross-linking studies on the McjD dimer in inside-out membrane vesicles of E. coli confirmed the presence of the occluded state. We therefore propose that the outward-occluded state represents a transition intermediate between the outward-open and inward-open conformation of ABC exporters.
Journal Keywords: Amino; Bacteriocins; Crystallography; X-Ray; Escherichia; Mutagenesis; Site-Directed; Mutation; Missense; Protein; Quaternary; Protein; Secondary; Protein; Tertiary
Diamond Keywords: Bacteria
Subject Areas:
Biology and Bio-materials,
Chemistry
Instruments:
I02-Macromolecular Crystallography
Added On:
16/09/2014 09:23
Documents:
PNAS-2014-Choudhury-9145-50.pdf
Discipline Tags:
Pathogens
Infectious Diseases
Health & Wellbeing
Biochemistry
Chemistry
Structural biology
Life Sciences & Biotech
Technical Tags:
Diffraction
Macromolecular Crystallography (MX)