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An on-demand, drop-on-drop method for studying enzyme catalysis by serial crystallography

Author

Listed:
  • Agata Butryn

    (Harwell Science and Innovation Campus
    Rutherford Appleton Laboratory)

  • Philipp S. Simon

    (Lawrence Berkeley National Laboratory)

  • Pierre Aller

    (Harwell Science and Innovation Campus
    Rutherford Appleton Laboratory)

  • Philip Hinchliffe

    (University Walk)

  • Ramzi N. Massad

    (Lawrence Berkeley National Laboratory)

  • Gabriel Leen

    (Rathmacullig West
    University of Limerick)

  • Catherine L. Tooke

    (University Walk)

  • Isabel Bogacz

    (Lawrence Berkeley National Laboratory)

  • In-Sik Kim

    (Lawrence Berkeley National Laboratory)

  • Asmit Bhowmick

    (Lawrence Berkeley National Laboratory)

  • Aaron S. Brewster

    (Lawrence Berkeley National Laboratory)

  • Nicholas E. Devenish

    (Harwell Science and Innovation Campus)

  • Jürgen Brem

    (University of Oxford)

  • Jos J. A. G. Kamps

    (University of Oxford
    Harwell Science and Innovation Campus)

  • Pauline A. Lang

    (University of Oxford)

  • Patrick Rabe

    (University of Oxford)

  • Danny Axford

    (Harwell Science and Innovation Campus)

  • John H. Beale

    (Harwell Science and Innovation Campus
    Paul Scherrer Institut)

  • Bradley Davy

    (Harwell Science and Innovation Campus
    University of Leeds)

  • Ali Ebrahim

    (Harwell Science and Innovation Campus)

  • Julien Orlans

    (Harwell Science and Innovation Campus
    University of Lyon)

  • Selina L. S. Storm

    (Harwell Science and Innovation Campus
    Hamburg Outstation c/o DESY)

  • Tiankun Zhou

    (Harwell Science and Innovation Campus
    Rutherford Appleton Laboratory)

  • Shigeki Owada

    (RIKEN SPring-8 Center
    Japan Synchrotron Radiation Research Institute)

  • Rie Tanaka

    (RIKEN SPring-8 Center
    Kyoto University)

  • Kensuke Tono

    (RIKEN SPring-8 Center
    Japan Synchrotron Radiation Research Institute)

  • Gwyndaf Evans

    (Harwell Science and Innovation Campus)

  • Robin L. Owen

    (Harwell Science and Innovation Campus)

  • Frances A. Houle

    (Lawrence Berkeley National Laboratory)

  • Nicholas K. Sauter

    (Lawrence Berkeley National Laboratory)

  • Christopher J. Schofield

    (University of Oxford)

  • James Spencer

    (University Walk)

  • Vittal K. Yachandra

    (Lawrence Berkeley National Laboratory)

  • Junko Yano

    (Lawrence Berkeley National Laboratory)

  • Jan F. Kern

    (Lawrence Berkeley National Laboratory)

  • Allen M. Orville

    (Harwell Science and Innovation Campus
    Rutherford Appleton Laboratory)

Abstract

Serial femtosecond crystallography has opened up many new opportunities in structural biology. In recent years, several approaches employing light-inducible systems have emerged to enable time-resolved experiments that reveal protein dynamics at high atomic and temporal resolutions. However, very few enzymes are light-dependent, whereas macromolecules requiring ligand diffusion into an active site are ubiquitous. In this work we present a drop-on-drop sample delivery system that enables the study of enzyme-catalyzed reactions in microcrystal slurries. The system delivers ligand solutions in bursts of multiple picoliter-sized drops on top of a larger crystal-containing drop inducing turbulent mixing and transports the mixture to the X-ray interaction region with temporal resolution. We demonstrate mixing using fluorescent dyes, numerical simulations and time-resolved serial femtosecond crystallography, which show rapid ligand diffusion through microdroplets. The drop-on-drop method has the potential to be widely applicable to serial crystallography studies, particularly of enzyme reactions with small molecule substrates.

Suggested Citation

  • Agata Butryn & Philipp S. Simon & Pierre Aller & Philip Hinchliffe & Ramzi N. Massad & Gabriel Leen & Catherine L. Tooke & Isabel Bogacz & In-Sik Kim & Asmit Bhowmick & Aaron S. Brewster & Nicholas E., 2021. "An on-demand, drop-on-drop method for studying enzyme catalysis by serial crystallography," Nature Communications, Nature, vol. 12(1), pages 1-7, December.
    • Handle: RePEc:nat:natcom:v:12:y:2021:i:1:d:10.1038_s41467-021-24757-7
    DOI: 10.1038/s41467-021-24757-7
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    Cited by:

    1. Basudev Maity & Mitsuo Shoji & Fangjia Luo & Takanori Nakane & Satoshi Abe & Shigeki Owada & Jungmin Kang & Kensuke Tono & Rie Tanaka & Thuc Toan Pham & Mariko Kojima & Yuki Hishikawa & Junko Tanaka &, 2024. "Real-time observation of a metal complex-driven reaction intermediate using a porous protein crystal and serial femtosecond crystallography," Nature Communications, Nature, vol. 15(1), pages 1-12, December.
    2. Maximilian Wranik & Tobias Weinert & Chavdar Slavov & Tiziana Masini & Antonia Furrer & Natacha Gaillard & Dario Gioia & Marco Ferrarotti & Daniel James & Hannah Glover & Melissa Carrillo & Demet Keki, 2023. "Watching the release of a photopharmacological drug from tubulin using time-resolved serial crystallography," Nature Communications, Nature, vol. 14(1), pages 1-12, December.

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