title: Low-dose X-ray radiation induces structural alterations in proteins.
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.
authors.affiliation: Université Grenoble Alpes, IBS, 38044 Grenoble, France.
authors.forename: Valentin
authors.initials: V
authors.lastname: Borshchevskiy
authors.affiliation: Université Grenoble Alpes, IBS, 38044 Grenoble, France.
authors.forename: Ekaterina
authors.initials: E
authors.lastname: Round
authors.affiliation: Laboratory for Advanced Studies of Membrane Proteins, Moscow Institute of Physics and Technology, Dolgoprudniy 141700, Russian Federation.
authors.forename: Ivan
authors.initials: I
authors.lastname: Erofeev
authors.affiliation: Université Grenoble Alpes, IBS, 38044 Grenoble, France.
authors.forename: Martin
authors.initials: M
authors.lastname: Weik
authors.affiliation: Institute of Complex Systems (ICS), ICS-6: Structural Biochemistry, Research Centre Jülich, 52425 Jülich, Germany.
authors.forename: Andrii
authors.initials: A
authors.lastname: Ishchenko
authors.affiliation: Université Grenoble Alpes, IBS, 38044 Grenoble, France.
authors.forename: Ivan
authors.initials: I
authors.lastname: Gushchin
authors.affiliation: Laboratory for Structural Biology of GPCRs, Moscow Institute of Physics and Technology, Dolgoprudniy 141700, Russian Federation.
authors.forename: Alexey
authors.initials: A
authors.lastname: Mishin
authors.affiliation: Institute of Complex Systems (ICS), ICS-6: Structural Biochemistry, Research Centre Jülich, 52425 Jülich, Germany.
authors.forename: Dieter
authors.initials: D
authors.lastname: Willbold
authors.affiliation: Laboratory for Advanced Studies of Membrane Proteins, Moscow Institute of Physics and Technology, Dolgoprudniy 141700, Russian Federation.
authors.forename: Georg
authors.initials: G
authors.lastname: Büldt
authors.affiliation: Université Grenoble Alpes, IBS, 38044 Grenoble, France.
authors.forename: Valentin
authors.initials: V
authors.lastname: Gordeliy
mesh_terms: D001436:Bacteriorhodopsins / Q000737:chemistry*; D020134:Catalytic Domain; D018360:Crystallography, X-Ray / Q000379:methods*; D004307:Dose-Response Relationship, Radiation; D005583:Fourier Analysis; D008958:Models, Molecular; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry* / Q000528:radiation effects*; D014965:X-Rays
chemical_list: D011506:Proteins; D001436:Bacteriorhodopsins
keywords: bacteriorhodopsin; in meso crystallization; intermediate states; orange species; radiation damage