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Title: In cellulo serial crystallography of alcohol oxidase crystals inside yeast cells

The possibility of using femtosecond pulses from an X-ray free-electron laser to collect diffraction data from protein crystals formed in their native cellular organelle has been explored. X-ray diffraction of submicrometre-sized alcohol oxidase crystals formed in peroxisomes within cells of genetically modified variants of the methylotrophic yeast Hansenula polymorpha is reported and characterized. Furthermore, the observations are supported by synchrotron radiation-based powder diffraction data and electron microscopy. Based on these findings, the concept of in cellulo serial crystallography on protein targets imported into yeast peroxisomes without the need for protein purification as a requirement for subsequent crystallization is outlined.
Authors:
 [1] ;  [2] ;  [3] ;  [2] ;  [2] ;  [2] ;  [2] ;  [4] ;  [5] ;  [6]
  1. Deutsches Elektronen-Synchrotron (DESY), Hamburg (Germany); European Molecular Biology Lab. (EMBL), Heidelberg (Germany)
  2. Deutsches Elektronen-Synchrotron (DESY), Hamburg (Germany)
  3. Univ. of Groningen, Groningen (The Netherlands)
  4. SLAC National Accelerator Lab., Menlo Park, CA (United States)
  5. Deutsches Elektronen-Synchrotron (DESY), Hamburg (Germany); Univ. of Hamburg, Hamburg (Germany); Center for Ultrafast Imaging, Hamburg (Germany)
  6. Deutsches Elektronen-Synchrotron (DESY), Hamburg (Germany); Univ. Medical Center Hamburg-Eppendorf, Hamburg (Germany)
Publication Date:
OSTI Identifier:
1244994
Grant/Contract Number:
AC02-76SF00515
Type:
Accepted Manuscript
Journal Name:
IUCrJ
Additional Journal Information:
Journal Volume: 3; Journal Issue: 2; Journal ID: ISSN 2052-2525
Publisher:
International Union of Crystallography
Research Org:
SLAC National Accelerator Laboratory (SLAC), Menlo Park, CA (United States)
Sponsoring Org:
USDOE Office of Science (SC)
Country of Publication:
United States
Language:
English
Subject:
37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY protein structure; X-ray crystallography; femtosecond studies; nanocrystals; free-electron laser