Cryotrapping peroxide in the active site of human mitochondrial manganese superoxide dismutase crystals for neutron diffraction

Jahaun Azadmanesh, William E. Lutz, Leighton Coates, Kevin L. Weiss, Gloria E.O. Borgstahl

Research output: Contribution to journalArticlepeer-review

1 Scopus citations

Abstract

Structurally identifying the enzymatic intermediates of redox proteins has been elusive due to difficulty in resolving the H atoms involved in catalysis and the susceptibility of ligand complexes to photoreduction from X-rays. Cryotrapping ligands for neutron protein crystallography combines two powerful tools that offer the advantage of directly identifying hydrogen positions in redox-enzyme intermediates without radiolytic perturbation of metal-containing active sites. However, translating cryogenic techniques from X-ray to neutron crystallography is not straightforward due to the large crystal volumes and long data-collection times. Here, methods have been developed to visualize the evasive peroxo complex of manganese superoxide dismutase (MnSOD) so that all atoms, including H atoms, could be visualized. The subsequent cryocooling and ligand-trapping methods resulted in neutron data collection to 2.30 Å resolution. The P6122 crystal form of MnSOD is challenging because it has some of the largest unit-cell dimensions (a = b = 77.8, c = 236.8 Å) ever studied using high-resolution cryo-neutron crystallography. The resulting neutron diffraction data permitted the visualization of a dioxygen species bound to the MnSOD active-site metal that was indicative of successful cryotrapping.

Original languageEnglish (US)
Pages (from-to)8-16
Number of pages9
JournalActa Crystallographica Section F: Structural Biology Communications
Volume78
DOIs
StatePublished - Jan 1 2022

Keywords

  • Cryotrapping
  • Human manganese superoxide dismutase
  • Large unit cell
  • Neutron diffraction
  • Peroxide

ASJC Scopus subject areas

  • Biophysics
  • Structural Biology
  • Biochemistry
  • Genetics
  • Condensed Matter Physics

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