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Insights into receptor structure and dynamics at the surface of living cells

Author

Listed:
  • Frederik Steiert

    (Max Planck Institute of Biochemistry
    Technical University Munich)

  • Peter Schultz

    (Max Planck Institute of Biochemistry)

  • Siegfried Höfinger

    (VSC Research Center, TU Wien
    Michigan Technological University)

  • Thomas D. Müller

    (Lehrstuhl für Molekulare Pflanzenphysiologie und Biophysik - Botanik I)

  • Petra Schwille

    (Max Planck Institute of Biochemistry)

  • Thomas Weidemann

    (Max Planck Institute of Biochemistry)

Abstract

Evaluating protein structures in living cells remains a challenge. Here, we investigate Interleukin-4 receptor alpha (IL-4Rα) into which the non-canonical amino acid bicyclo[6.1.0]nonyne-lysine (BCNK) is incorporated by genetic code expansion. Bioorthogonal click labeling is performed with tetrazine-conjugated dyes. To quantify the reaction yield in situ, we develop brightness-calibrated ratiometric imaging, a protocol where fluorescent signals in confocal multi-color images are ascribed to local concentrations. Screening receptor mutants bearing BCNK in the extracellular domain uncovered site-specific variations of both click efficiency and Interleukin-4 binding affinity, indicating subtle well-defined structural perturbations. Molecular dynamics and continuum electrostatics calculations suggest solvent polarization to determine site-specific variations of BCNK reactivity. Strikingly, signatures of differential click efficiency, measured for IL-4Rα in ligand-bound and free form, mirror sub-angstrom deformations of the protein backbone at corresponding locations. Thus, click efficiency by itself represents a remarkably informative readout linked to protein structure and dynamics in the native plasma membrane.

Suggested Citation

  • Frederik Steiert & Peter Schultz & Siegfried Höfinger & Thomas D. Müller & Petra Schwille & Thomas Weidemann, 2023. "Insights into receptor structure and dynamics at the surface of living cells," Nature Communications, Nature, vol. 14(1), pages 1-15, December.
    • Handle: RePEc:nat:natcom:v:14:y:2023:i:1:d:10.1038_s41467-023-37284-4
    DOI: 10.1038/s41467-023-37284-4
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    References listed on IDEAS

    as
    1. Martin Baumdick & Márton Gelléri & Chayasith Uttamapinant & Václav Beránek & Jason W. Chin & Philippe I. H. Bastiaens, 2018. "A conformational sensor based on genetic code expansion reveals an autocatalytic component in EGFR activation," Nature Communications, Nature, vol. 9(1), pages 1-13, December.
    2. Diogo Bessa-Neto & Gerti Beliu & Alexander Kuhlemann & Valeria Pecoraro & Sören Doose & Natacha Retailleau & Nicolas Chevrier & David Perrais & Markus Sauer & Daniel Choquet, 2021. "Bioorthogonal labeling of transmembrane proteins with non-canonical amino acids unveils masked epitopes in live neurons," Nature Communications, Nature, vol. 12(1), pages 1-16, December.
    3. David Richter & Ignacio Moraga & Hauke Winkelmann & Oliver Birkholz & Stephan Wilmes & Markos Schulte & Michael Kraich & Hella Kenneweg & Oliver Beutel & Philipp Selenschik & Dirk Paterok & Martynas G, 2017. "Ligand-induced type II interleukin-4 receptor dimers are sustained by rapid re-association within plasma membrane microcompartments," Nature Communications, Nature, vol. 8(1), pages 1-15, December.
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