On-chip crystallization for serial crystallography experiments and on-chip ligand-binding studies

Forskningsoutput: TidskriftsbidragArtikel i vetenskaplig tidskrift

Abstract

Efficient and reliable sample delivery has remained one of the bottlenecks for serial crystallography experiments. Compared with other methods, fixed-target sample delivery offers the advantage of significantly reduced sample consumption and shorter data collection times owing to higher hit rates. Here, a new method of on-chip crystallization is reported which allows the efficient and reproducible growth of large numbers of protein crystals directly on micro-patterned silicon chips for in-situ serial crystallography experiments. Crystals are grown by sitting-drop vapor diffusion and previously established crystallization conditions can be directly applied. By reducing the number of crystal-handling steps, the method is particularly well suited for sensitive crystal systems. Excessive mother liquor can be efficiently removed from the crystals by blotting, and no sealing of the fixed-target sample holders is required to prevent the crystals from dehydrating. As a consequence, 'naked' crystals are obtained on the chip, resulting in very low background scattering levels and making the crystals highly accessible for external manipulation such as the application of ligand solutions. Serial diffraction experiments carried out at cryogenic temperatures at a synchrotron and at room temperature at an X-ray free-electron laser yielded high-quality X-ray structures of the human membrane protein aquaporin 2 and two new ligand-bound structures of thermolysin and the human kinase DRAK2. The results highlight the applicability of the method for future high-throughput on-chip screening of pharmaceutical compounds.

Detaljer

Författare
  • Julia Lieske
  • Maximilian Cerv
  • Dana Komadina
  • Janine Fischer
  • Miriam Barthelmess
  • Pontus Fischer
  • Tim Pakendorf
  • Oleksandr Yefanov
  • Valerio Mariani
  • Thomas Seine
  • Breyan H. Ross
  • Eva Crosas
  • Olga Lorbeer
  • Anja Burkhardt
  • Thomas J. Lane
  • Sebastian Guenther
  • Julian Bergtholdt
  • Silvan Schoen
  • Henry N. Chapman
  • Alke Meents
Enheter & grupper
Externa organisationer
  • German Electron Synchrotron (DESY)
  • European Molecular Biology Laboratory Hamburg
  • Max Planck Institute of Biochemistry
  • SLAC National Accelerator Laboratory
  • University of Hamburg
Forskningsområden

Ämnesklassifikation (UKÄ) – OBLIGATORISK

  • Strukturbiologi
  • Biokemi och molekylärbiologi

Nyckelord

Originalspråkengelska
Sidor (från-till)714-728
Antal sidor15
TidskriftIUCrJ
Volym6
Utgivningsnummer4
StatusPublished - 2019 jul
PublikationskategoriForskning
Peer review utfördJa