X-ray Free Electron Laser Determination of Crystal Structures of Dark and Light States of a Reversibly Photoswitching Fluorescent Protein at Room Temperature.
Int J Mol Sci
; 18(9)2017 Sep 07.
Article
em En
| MEDLINE
| ID: mdl-28880248
ABSTRACT
The photochromic fluorescent protein Skylan-NS (Nonlinear Structured illumination variant mEos3.1H62L) is a reversibly photoswitchable fluorescent protein which has an unilluminated/ground state with an anionic and cis chromophore conformation and high fluorescence quantum yield. Photo-conversion with illumination at 515 nm generates a meta-stable intermediate with neutral trans-chromophore structure that has a 4 h lifetime. We present X-ray crystal structures of the cis (on) state at 1.9 Angstrom resolution and the trans (off) state at a limiting resolution of 1.55 Angstrom from serial femtosecond crystallography experiments conducted at SPring-8 Angstrom Compact Free Electron Laser (SACLA) at 7.0 keV and 10.5 keV, and at Linac Coherent Light Source (LCLS) at 9.5 keV. We present a comparison of the data reduction and structure determination statistics for the two facilities which differ in flux, beam characteristics and detector technologies. Furthermore, a comparison of droplet on demand, grease injection and Gas Dynamic Virtual Nozzle (GDVN) injection shows no significant differences in limiting resolution. The photoconversion of the on- to the off-state includes both internal and surface exposed protein structural changes, occurring in regions that lack crystal contacts in the orthorhombic crystal form.
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Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Cristalografia por Raios X
/
Lasers
/
Proteínas Luminescentes
Idioma:
En
Revista:
Int J Mol Sci
Ano de publicação:
2017
Tipo de documento:
Article
País de afiliação:
Reino Unido