Microcrystal preparation for serial femtosecond X-ray crystallography of bacterial copper amine oxidase.
Acta Crystallogr F Struct Biol Commun
; 77(Pt 10): 356-363, 2021 Oct 01.
Article
em En
| MEDLINE
| ID: mdl-34605440
ABSTRACT
Recent advances in serial femtosecond X-ray crystallography (SFX) using X-ray free-electron lasers have paved the way for determining radiation-damage-free protein structures under nonfreezing conditions. However, the large-scale preparation of high-quality microcrystals of uniform size is a prerequisite for SFX, and this has been a barrier to its widespread application. Here, a convenient method for preparing high-quality microcrystals of a bacterial quinoprotein enzyme, copper amine oxidase from Arthrobacter globiformis, is reported. The method consists of the mechanical crushing of large crystals (5-15â
mm3), seeding the crushed crystals into the enzyme solution and standing for 1â
h at an ambient temperature of â¼26°C, leading to the rapid formation of microcrystals with a uniform size of 3-5â
µm. The microcrystals diffracted X-rays to a resolution beyond 2.0â
Å in SFX measurements at the SPring-8 Angstrom Compact Free Electron Laser facility. The damage-free structure determined at 2.2â
Å resolution was essentially identical to that determined previously by cryogenic crystallography using synchrotron X-ray radiation.
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MEDLINE
Assunto principal:
Arthrobacter
/
Amina Oxidase (contendo Cobre)
/
Síncrotrons
Idioma:
En
Ano de publicação:
2021
Tipo de documento:
Article