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Two states of a light-sensitive membrane protein captured at room temperature using thin-film sample mounts
Room-temperature diffraction methods are highly desirable for dynamic studies of biological macromolecules, since they allow high-resolution structural data to be collected as proteins undergo conformational changes. For crystals grown in lipidic cubic phase (LCP), an extruder is commonly used to pa...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
International Union of Crystallography
2022
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8725165/ https://www.ncbi.nlm.nih.gov/pubmed/34981761 http://dx.doi.org/10.1107/S2059798321011220 |
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author | Axford, Danny Judge, Peter J. Bada Juarez, Juan F. Kwan, Tristan O. C. Birch, James Vinals, Javier Watts, Anthony Moraes, Isabel |
author_facet | Axford, Danny Judge, Peter J. Bada Juarez, Juan F. Kwan, Tristan O. C. Birch, James Vinals, Javier Watts, Anthony Moraes, Isabel |
author_sort | Axford, Danny |
collection | PubMed |
description | Room-temperature diffraction methods are highly desirable for dynamic studies of biological macromolecules, since they allow high-resolution structural data to be collected as proteins undergo conformational changes. For crystals grown in lipidic cubic phase (LCP), an extruder is commonly used to pass a stream of microcrystals through the X-ray beam; however, the sample quantities required for this method may be difficult to produce for many membrane proteins. A more sample-efficient environment was created using two layers of low X-ray transmittance polymer films to mount crystals of the archaerhodopsin-3 (AR3) photoreceptor and room-temperature diffraction data were acquired. By using transparent and opaque polymer films, two structures, one corresponding to the desensitized, dark-adapted (DA) state and the other to the ground or light-adapted (LA) state, were solved to better than 1.9 Å resolution. All of the key structural features of AR3 were resolved, including the retinal chromophore, which is present as the 13-cis isomer in the DA state and as the all-trans isomer in the LA state. The film-sandwich sample environment enables diffraction data to be recorded at room temperature in both illuminated and dark conditions, which more closely approximate those in vivo. This simple approach is applicable to a wide range of membrane proteins crystallized in LCP and light-sensitive samples in general at synchrotron and laboratory X-ray sources. |
format | Online Article Text |
id | pubmed-8725165 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | International Union of Crystallography |
record_format | MEDLINE/PubMed |
spelling | pubmed-87251652022-01-06 Two states of a light-sensitive membrane protein captured at room temperature using thin-film sample mounts Axford, Danny Judge, Peter J. Bada Juarez, Juan F. Kwan, Tristan O. C. Birch, James Vinals, Javier Watts, Anthony Moraes, Isabel Acta Crystallogr D Struct Biol Research Papers Room-temperature diffraction methods are highly desirable for dynamic studies of biological macromolecules, since they allow high-resolution structural data to be collected as proteins undergo conformational changes. For crystals grown in lipidic cubic phase (LCP), an extruder is commonly used to pass a stream of microcrystals through the X-ray beam; however, the sample quantities required for this method may be difficult to produce for many membrane proteins. A more sample-efficient environment was created using two layers of low X-ray transmittance polymer films to mount crystals of the archaerhodopsin-3 (AR3) photoreceptor and room-temperature diffraction data were acquired. By using transparent and opaque polymer films, two structures, one corresponding to the desensitized, dark-adapted (DA) state and the other to the ground or light-adapted (LA) state, were solved to better than 1.9 Å resolution. All of the key structural features of AR3 were resolved, including the retinal chromophore, which is present as the 13-cis isomer in the DA state and as the all-trans isomer in the LA state. The film-sandwich sample environment enables diffraction data to be recorded at room temperature in both illuminated and dark conditions, which more closely approximate those in vivo. This simple approach is applicable to a wide range of membrane proteins crystallized in LCP and light-sensitive samples in general at synchrotron and laboratory X-ray sources. International Union of Crystallography 2022-01-01 /pmc/articles/PMC8725165/ /pubmed/34981761 http://dx.doi.org/10.1107/S2059798321011220 Text en © Danny Axford et al. 2022 https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution (CC-BY) Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original authors and source are cited. |
spellingShingle | Research Papers Axford, Danny Judge, Peter J. Bada Juarez, Juan F. Kwan, Tristan O. C. Birch, James Vinals, Javier Watts, Anthony Moraes, Isabel Two states of a light-sensitive membrane protein captured at room temperature using thin-film sample mounts |
title | Two states of a light-sensitive membrane protein captured at room temperature using thin-film sample mounts |
title_full | Two states of a light-sensitive membrane protein captured at room temperature using thin-film sample mounts |
title_fullStr | Two states of a light-sensitive membrane protein captured at room temperature using thin-film sample mounts |
title_full_unstemmed | Two states of a light-sensitive membrane protein captured at room temperature using thin-film sample mounts |
title_short | Two states of a light-sensitive membrane protein captured at room temperature using thin-film sample mounts |
title_sort | two states of a light-sensitive membrane protein captured at room temperature using thin-film sample mounts |
topic | Research Papers |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8725165/ https://www.ncbi.nlm.nih.gov/pubmed/34981761 http://dx.doi.org/10.1107/S2059798321011220 |
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