De-novo macrocyclic peptides dissect energy coupling of a heterodimeric ABC transporter by multimode allosteric inhibition

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Abstract

ATP-binding cassette (ABC) transporters constitute the largest family of primary active transporters involved in a multitude of physiological processes and human diseases. Despite considerable efforts, it remains unclear how ABC transporters harness the chemical energy of ATP to drive substrate transport across cell membranes. Here, by random nonstandard peptide integrated discovery (RaPID), we leveraged combinatorial macrocyclic peptides that target a heterodimeric ABC transport complex and explore fundamental principles of the substrate translocation cycle. High-affinity peptidic macrocycles bind conformationally selective and display potent multimode inhibitory effects. The macrocycles block the transporter either before or after unidirectional substrate export along a single conformational switch induced by ATP binding. Our study reveals mechanistic principles of ATP binding, conformational switching, and energy transduction for substrate transport of ABC export systems. We highlight the potential of de novo macrocycles as effective inhibitors for membrane proteins implicated in multidrug resistance, providing avenues for the next-generation of pharmaceuticals.

Data availability

All data denerated or analyzed during this study are included in the manuscript and support files. A source data file has been provided for Figure 1C (Sequencing Data), Figure 2-6, Figure 2-supplement figure 1,and Figure 5-supplement figure 1.

Article and author information

Author details

Erich Stefan

Institute of Biochemistry, Biocenter, Goethe University Frankfurt, Frankfurt, Germany

Competing interests
The authors declare that no competing interests exist.
Richard Obexer

Department of Chemistry, The University of Tokyo, Tokyo, Japan

Competing interests
The authors declare that no competing interests exist.
Susanne Hofmann

Institute of Biochemistry, Biocenter, Goethe University Frankfurt, Frankfurt, Germany

Competing interests
The authors declare that no competing interests exist.
Khanh Vu Huu

3Institute of Physical and Theoretical Chemistry, Goethe University Frankfurt, Frankfurt, Germany

Competing interests
The authors declare that no competing interests exist.
Yichao Huang

Department of Chemistry, The University of Tokyo, Tokyo, Japan

Competing interests
The authors declare that no competing interests exist.
Nina Morgner

Institute of Physical and Theoretical Chemistry, Goethe University Frankfurt, Frankfurt, Germany

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-1872-490X
Hiroaki Suga

Department of Chemistry, The University of Tokyo, Tokyo, Japan

For correspondence
hsuga@chem.su-tokyo.ac.jp

Competing interests
The authors declare that no competing interests exist.
Robert Tampé

Institute of Biochemistry, Biocenter, Goethe University Frankfurt, Frankfurt, Germany

For correspondence
tampe@em.uni-frankfurt.de

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-0403-2160

Funding

Deutsche Forschungsgemeinschaft (TA 157/12-1)

Robert Tampé

Deutsche Forschungsgemeinschaft (CRC 807/P16)

Robert Tampé

Deutsche Forschungsgemeinschaft (CRC 807/24)

Nina Morgner

European Research Council (798121)

Robert Tampé

JSPS Grants-in-Aid for Research Fellowship (P15333)

Richard Obexer

Japan Society for the Promotion of Science (JP20H05618)

Hiroaki Suga

Japan Society for the Promotion of Science (JP20am0101090)

Hiroaki Suga

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.