Structure and ligand binding in the putative anti-microbial peptide transporter protein, YejA

Bryony K Ackroyd, Eleanor J Dodson, Javeria Mehboob, Adam A Dowle, Gavin H Thomas*, Anthony J Wilkinson*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review


YejABEF is an ATP-binding cassette transporter that is implicated in the sensitivity of Escherichia coli to anti-microbial peptides, the best-characterized example being microcin C, a peptide-nucleotide antibiotic that targets aspartyl-tRNA synthetase. Here the structure of the extracellular solute binding protein, YejA, has been determined, revealing an oligopeptide-binding protein fold enclosing a ligand-binding pocket larger than those of other peptide-binding proteins of known structure. Prominent electron density in this cavity defines an undecapeptide sequence LGEPRYAFNFN, an observation that is confirmed by mass spectrometry. In the structure, the peptide interactions with the protein are mediated by main chain hydrogen bonds with the exception of Arg5 whose guanidinium side chain makes a set of defining polar interactions with four YejA residues. More detailed characterization of purified recombinant YejA, by a combination of ESI and MALDI-mass spectrometry as well as thermal shift assays, reveals a set of YejA complexes containing overlapping peptides 10–19 residues in length. All contain the sequence LGEPRYAFN. Curiously, these peptides correspond to residues 8–26 of the mature YejA protein, which belong to a unique N-terminal extension that distinguishes YejA from other cluster C oligopeptide binding proteins of known structure. This 35-residue extension is well-ordered and packs across the surface of the protein. The undecapeptide ligand occupies only a fraction of the enclosed pocket volume suggesting the possibility that much larger peptides or peptide conjugates could be accommodated, though thermal shift assays of YejA binding to antimicrobial peptides and peptides unrelated to LGEPRYAFNFN have not provided evidence of binding. While the physiological significance of this ‘auto-binding’ is not clear, the experimental data suggest that it is not an artefact of the crystallization process and that it may have a function in the sensing of periplasmic or membrane stress.

Original languageEnglish
Article number001430
Number of pages16
JournalMicrobiology (Reading, England)
Issue number2
Publication statusPublished - 9 Feb 2024

Bibliographical note

Funding Information:
We are grateful to Sam Hart and Johan Turkenburg for help and advice with diffraction data collection and processing and to Andrew Leech and Jared Cartwright of the Bioscience Technology Facility at York for technical assistance. The York Centre of Excellence in Mass Spectrometry was created thanks to a major capital investment through Science City York, supported by Yorkshire Forward with funds from the Northern Way Initiative, and subsequent support from EPSRC (EP/K039660/1; EP/M028127/1). We thank the Diamond Light Source for access to beamlines i04 (proposal number mx-13587) that contributed to the results presented here.

Funding Information:
This project was supported by the Wellcome Trust through a studentship awarded to BKA as part of the University of York PhD Programme CIDCATS (105501/Z/14/Z).

Publisher Copyright:
© 2024 The Authors.


  • Ligands
  • Peptides/metabolism
  • ATP-Binding Cassette Transporters/genetics
  • Oligopeptides
  • Escherichia coli/metabolism
  • Protein Binding
  • Membrane Transport Proteins

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