Structural and mechanistic analysis of a tripartite ATP-independent periplasmic TRAP transporter

Martin F Peter, Jan A Ruland, Peer Depping, Niels Schneberger, Emmanuele Severi, Jonas Moecking, Karl Gatterdam, Sarah Tindall, Alexandre Durand, Veronika Heinz, Jan Peter Siebrasse, Paul-Albert Koenig, Matthias Geyer, Christine Ziegler, Ulrich Kubitscheck, Gavin H Thomas, Gregor Hagelueken

Research output: Contribution to journalArticlepeer-review

Abstract

Tripartite ATP-independent periplasmic (TRAP) transporters are found widely in bacteria and archaea and consist of three structural domains, a soluble substrate-binding protein (P-domain), and two transmembrane domains (Q- and M-domains). HiSiaPQM and its homologs are TRAP transporters for sialic acid and are essential for host colonization by pathogenic bacteria. Here, we reconstitute HiSiaQM into lipid nanodiscs and use cryo-EM to reveal the structure of a TRAP transporter. It is composed of 16 transmembrane helices that are unexpectedly structurally related to multimeric elevator-type transporters. The idiosyncratic Q-domain of TRAP transporters enables the formation of a monomeric elevator architecture. A model of the tripartite PQM complex is experimentally validated and reveals the coupling of the substrate-binding protein to the transporter domains. We use single-molecule total internal reflection fluorescence (TIRF) microscopy in solid-supported lipid bilayers and surface plasmon resonance to study the formation of the tripartite complex and to investigate the impact of interface mutants. Furthermore, we characterize high-affinity single variable domains on heavy chain (VHH) antibodies that bind to the periplasmic side of HiSiaQM and inhibit sialic acid uptake, providing insight into how TRAP transporter function might be inhibited in vivo.

Original languageEnglish
Article number4471
Pages (from-to)4471
JournalNature Communications
Volume13
Issue number1
Early online date4 Aug 2022
DOIs
Publication statusPublished - 4 Aug 2022

Bibliographical note

© 2022. The Author(s). Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give
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Funding Information:
Open Access funding enabled and organized by Projekt DEAL.

Keywords

  • Adenosine Triphosphate/metabolism
  • Archaea/metabolism
  • Bacteria/metabolism
  • Bacterial Proteins/metabolism
  • Carrier Proteins/metabolism
  • Membrane Transport Proteins/metabolism
  • N-Acetylneuraminic Acid/metabolism

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