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Acta Crystallogr D Biol Crystallogr. 2014 Oct;70(Pt 10):2675-85. doi: 10.1107/S1399004714017295. Epub 2014 Sep 27.

Low-dose X-ray radiation induces structural alterations in proteins.

Author information

1
Université Grenoble Alpes, IBS, 38044 Grenoble, France.
2
Laboratory for Advanced Studies of Membrane Proteins, Moscow Institute of Physics and Technology, Dolgoprudniy 141700, Russian Federation.
3
Institute of Complex Systems (ICS), ICS-6: Structural Biochemistry, Research Centre Jülich, 52425 Jülich, Germany.
4
Laboratory for Structural Biology of GPCRs, Moscow Institute of Physics and Technology, Dolgoprudniy 141700, Russian Federation.

Abstract

X-ray-radiation-induced alterations to protein structures are still a severe problem in macromolecular crystallography. One way to avoid the influence of radiation damage is to reduce the X-ray dose absorbed by the crystal during data collection. However, here it is demonstrated using the example of the membrane protein bacteriorhodopsin (bR) that even a low dose of less than 0.06 MGy may induce structural alterations in proteins. This dose is about 500 times smaller than the experimental dose limit which should ideally not be exceeded per data set (i.e. 30 MGy) and 20 times smaller than previously detected specific radiation damage at the bR active site. To date, it is the lowest dose at which radiation modification of a protein structure has been described. Complementary use was made of high-resolution X-ray crystallography and online microspectrophotometry to quantitatively study low-dose X-ray-induced changes. It is shown that structural changes of the protein correlate with the spectroscopically observed formation of the so-called bR orange species. Evidence is provided for structural modifications taking place at the protein active site that should be taken into account in crystallographic studies which aim to elucidate the molecular mechanisms of bR function.

KEYWORDS:

bacteriorhodopsin; in meso crystallization; intermediate states; orange species; radiation damage

PMID:
25286851
DOI:
10.1107/S1399004714017295
[Indexed for MEDLINE]

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