X-ray-Radiation-Induced Changes in Bacteriorhodopsin Structure

Borshchevskiy, Valentin; Round, Ekaterina; Попов, А. Н.; Büldt, Georg; Gordeliy, Valentin

doi:10.1016/j.jmb.2011.04.038

Cited by 42 publications

(54 citation statements)

References 57 publications

(134 reference statements)

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“…In order to compare the influence of X-rays at 15 K with that at 100 K, a series of diffraction datasets were successively collected from a single bR crystal at each temperature (Supplementary Tables 1 and 2). At 100 K, the difference Fourier maps showed many positive and negative peaks, especially around the proton path, as previously reported 13,14 (Fig. 1a).…”

Section: Resultssupporting

confidence: 86%

X-ray structure analysis of bacteriorhodopsin at 1.3 Å resolution

Hasegawa

Jonotsuka

Miki

et al. 2018

Sci Rep

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Bacteriorhodopsin (bR) of Halobacterium salinarum is a membrane protein that acts as a light-driven proton pump. bR and its homologues have recently been utilized in optogenetics and other applications. Although the structures of those have been reported so far, the resolutions are not sufficient for elucidation of the intrinsic structural features critical to the color tuning and ion pumping properties. Here we report the accurate crystallographic analysis of bR in the ground state. The influence of X-rays was suppressed by collecting the data under a low irradiation dose at 15 K. Consequently, individual atoms could be separately observed in the electron density map at better than 1.3 Å resolution. Residues from Thr5 to Ala233 were continuously constructed in the model. The twist of the retinal polyene was determined to be different from those in the previous models. Two conformations were observed for the proton release region. We discuss the meaning of these fine structural features.

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Section: Resultssupporting

confidence: 86%

X-ray structure analysis of bacteriorhodopsin at 1.3 Å resolution

Hasegawa

Jonotsuka

Miki

et al. 2018

Sci Rep

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“…In particular, the cluster formed by Asp82, Asp85 and three water molecules (W401, W402 and W406) are crucial as the primary proton acceptor, ∼40 μs after the light-driven retinal isomerization from all- trans to 13- cis retinal. Light-induced conformational changes in this region, including rearrangements of water molecules are observed in cryo-trapped early photoactivation intermediates, but are also very similar to the changes induced by the X-ray radiation41. With continuous 2 F o − F c density for Asp82 and Asp85 and clear F o − F c difference density for the three water molecules, our SFX structure of bR resolves this region well (Fig.…”

Section: Resultssupporting

confidence: 68%

Lipidic cubic phase injector is a viable crystal delivery system for time-resolved serial crystallography

et al. 2016

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Serial femtosecond crystallography (SFX) using X-ray free-electron laser sources is an emerging method with considerable potential for time-resolved pump-probe experiments. Here we present a lipidic cubic phase SFX structure of the light-driven proton pump bacteriorhodopsin (bR) to 2.3 Å resolution and a method to investigate protein dynamics with modest sample requirement. Time-resolved SFX (TR-SFX) with a pump-probe delay of 1 ms yields difference Fourier maps compatible with the dark to M state transition of bR. Importantly, the method is very sample efficient and reduces sample consumption to about 1 mg per collected time point. Accumulation of M intermediate within the crystal lattice is confirmed by time-resolved visible absorption spectroscopy. This study provides an important step towards characterizing the complete photocycle dynamics of retinal proteins and demonstrates the feasibility of a sample efficient viscous medium jet for TR-SFX.

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“…In particular, the cluster formed by Asp82, Asp85 and three water molecules (W401, W402 and W406) are crucial as the primary proton acceptor, B40 ms after the light-driven retinal isomerization from all-trans to 13-cis retinal. Light-induced conformational changes in this region, including rearrangements of water molecules are observed in cryo-trapped early photoactivation intermediates, but are also very similar to the changes induced by the X-ray radiation 41 . With continuous 2F o À F c density for Asp82 and Asp85 and clear F o À F c difference density for the three water molecules, our SFX structure of bR resolves this region well (Fig.…”

Section: Resultsmentioning

confidence: 64%

“…At doses B1.2 MGy, far below the suggested maximal dose of 30 MGy for Cryo 53,54 , several X-ray-induced structural changes appear near the retinal-binding site. Since the spectroscopic properties of bR are strongly coupled to the isomerization of retinal and structural rearrangements of the surrounding amino acids and water molecules, X-ray exposure alone can lead to the formation of a bR species with active-like spectroscopic and structural characteristics 41,55 .…”

Section: Discussionmentioning

confidence: 99%