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

doi:10.1038/s41467-022-31907-y

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 journal › Article › peer-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 language	English
Article number	4471
Pages (from-to)	4471
Journal	Nature Communications
Volume	13
Issue number	1
Early online date	4 Aug 2022
DOIs	https://doi.org/10.1038/s41467-022-31907-y
Publication status	Published - 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
appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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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10.1038/s41467-022-31907-y

Structural and mechanistic analysis of a tripartite ATP-independent periplasmic TRAP transporter
© The Author(s) 2022
Final published version, 4.69 MBLicence: CC BY 4.0

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Peter, M. F., Ruland, J. A., Depping, P., Schneberger, N., Severi, E., Moecking, J., Gatterdam, K., Tindall, S., Durand, A., Heinz, V., Siebrasse, J. P., Koenig, P.-A., Geyer, M., Ziegler, C., Kubitscheck, U., Thomas, G. H., & Hagelueken, G. (2022). Structural and mechanistic analysis of a tripartite ATP-independent periplasmic TRAP transporter. Nature Communications, 13(1), 4471. Article 4471. https://doi.org/10.1038/s41467-022-31907-y

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Peter, MF, Ruland, JA, Depping, P, Schneberger, N, Severi, E, Moecking, J, Gatterdam, K, Tindall, S, Durand, A, Heinz, V, Siebrasse, JP, Koenig, P-A, Geyer, M, Ziegler, C, Kubitscheck, U, Thomas, GH & Hagelueken, G 2022, 'Structural and mechanistic analysis of a tripartite ATP-independent periplasmic TRAP transporter', Nature Communications, vol. 13, no. 1, 4471, pp. 4471. https://doi.org/10.1038/s41467-022-31907-y

TY - JOUR

T1 - Structural and mechanistic analysis of a tripartite ATP-independent periplasmic TRAP transporter

AU - Peter, Martin F

AU - Ruland, Jan A

AU - Depping, Peer

AU - Schneberger, Niels

AU - Severi, Emmanuele

AU - Moecking, Jonas

AU - Gatterdam, Karl

AU - Tindall, Sarah

AU - Durand, Alexandre

AU - Heinz, Veronika

AU - Siebrasse, Jan Peter

AU - Koenig, Paul-Albert

AU - Geyer, Matthias

AU - Ziegler, Christine

AU - Kubitscheck, Ulrich

AU - Thomas, Gavin H

AU - Hagelueken, Gregor

N1 - © 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 appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. Funding Information: Open Access funding enabled and organized by Projekt DEAL.

PY - 2022/8/4

Y1 - 2022/8/4

N2 - 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.

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KW - Adenosine Triphosphate/metabolism

KW - Archaea/metabolism

KW - Bacteria/metabolism

KW - Bacterial Proteins/metabolism

KW - Carrier Proteins/metabolism

KW - Membrane Transport Proteins/metabolism

KW - N-Acetylneuraminic Acid/metabolism

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DO - 10.1038/s41467-022-31907-y

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VL - 13

SP - 4471

JO - Nature Communications

JF - Nature Communications

IS - 1

M1 - 4471

ER -

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

Abstract

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Keywords

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