Cryoradiolytic reduction of crystalline heme proteins: analysis by UV-Vis spectroscopy and X-ray crystallography

Beitlich, T.; Kühnel, Karin; Schulze-Briese, Clemens; Shoeman, Robert L.; Schlichting, Ilme

doi:10.1107/s0909049506049806

Cited by 141 publications

(181 citation statements)

References 75 publications

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“…In another example, reduction of the haem iron by synchrotron radiation followed by transient warming to room temperature was used to generate and trap intermediate states in crystalline myoglobin (Hersleth et al, 2008). Research on other crystalline redox-sensitive proteins for which X-rayinduced reduction has been reported at temperatures close to 100 K (Berglund et al, 2002;Adam et al, 2004;Baxter et al, 2004;Mees et al, 2004;Roberts et al, 2005;Echalier et al, 2006;Beitlich et al, 2007;Kuhnel et al, 2007;Pearson et al, 2007;Hough et al, 2008) could benefit from temperaturecontrolled crystallography if structural information on intermediate states were of interest.…”

Section: Temperature-controlled Kinetic Cryocrystallography To Characmentioning

confidence: 99%

Temperature-dependent macromolecular X-ray crystallography

Weik

Colletier

2010

Acta Crystallogr D Biol Cryst

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X-ray crystallography provides structural details of biological macromolecules. Whereas routine data are collected close to 100 K in order to mitigate radiation damage, more exotic temperature-controlled experiments in a broader temperature range from 15 K to room temperature can provide both dynamical and structural insights. Here, the dynamical behaviour of crystalline macromolecules and their surrounding solvent as a function of cryo-temperature is reviewed. Experimental strategies of kinetic crystallography are discussed that have allowed the generation and trapping of macromolecular intermediate states by combining reaction initiation in the crystalline state with appropriate temperature profiles. A particular focus is on recruiting X-ray-induced changes for reaction initiation, thus unveiling useful aspects of radiation damage, which otherwise has to be minimized in macromolecular crystallography.

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Section: Temperature-controlled Kinetic Cryocrystallography To Characmentioning

confidence: 99%

Temperature-dependent macromolecular X-ray crystallography

Weik

Colletier

2010

Acta Crystallogr D Biol Cryst

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“…2F o -F c and F o -F c electron density maps display a diatomic molecule acting as the distal ligand. Because the heme iron atom can be photoreduced to the ferrous state because of the prolonged x-ray exposure (43)(44)(45), it was most likely in the ferrous form and coordinated by an oxygen molecule. The bond length between the oxygen molecule and heme iron is 2.35 Å.…”

Section: Efeb Binds Ppix and Enhances Its Fluorescence Intensity-mentioning

confidence: 99%

Crystal Structure and Biochemical Features of EfeB/YcdB from Escherichia coli O157

Wang

et al. 2011

Journal of Biological Chemistry

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EfeB/YcdB is a member of the dye-decolorizing peroxidase (DyP) protein family. A recent study has shown that this protein can extract iron from heme without breaking the tetrapyrrole ring. We report the crystal structure of EfeB from Escherichia coli O157 bound to heme at 1.95 Å resolution. The EfeB monomer contains two domains. The heme molecule is located in a large hydrophobic pocket in the C-terminal domain. A long loop connecting the two domains extensively interacts with the heme, which is a distinctive structural feature of EfeB homologues. A large tunnel formed by this loop and the ␤-sheet of C-terminal domain provides a potential cofactor/substrate binding site. Biochemical data show that the production of protoporphyrin IX (PPIX) is closely related to the peroxidation activity. The mutant D235N keeps nearly the same activity of guaiacol peroxidase as the wild-type protein, whereas the corresponding mutation in the classic DyP protein family completely abolished the peroxidation activity. These results suggest that EfeB is a unique member of the DyP protein family. In addition, dramatically enhanced fluorescence excitation and emission of EfeB-PPIX was observed, implying this protein may be used as a red color fluorescence marker.

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“…Photoreduction of protein-bound metal clusters was significantly slowed at temperatures below 100 K (25,27). However, photoreduction of metal clusters occurs at doses which are one to two orders of magnitude lower than the dose limit for global damage of around 30 MGy in cryocrystallography at 100 K (28,29).…”

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confidence: 99%

Origin and temperature dependence of radiation damage in biological samples at cryogenic temperatures

Meents

Gutmann

Wagner

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

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177

153

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Radiation damage is the major impediment for obtaining structural information from biological samples by using ionizing radiation such as x-rays or electrons. The knowledge of underlying processes especially at cryogenic temperatures is still fragmentary, and a consistent mechanism has not been found yet. By using a combination of single-crystal x-ray diffraction, small-angle scattering, and qualitative and quantitative radiolysis experiments, we show that hydrogen gas, formed inside the sample during irradiation, rather than intramolecular bond cleavage between non-hydrogen atoms, is mainly responsible for the loss of high-resolution information and contrast in diffraction experiments and microscopy. The experiments that are presented in this paper cover a temperature range between 5 and 160 K and reveal that the commonly used temperature in x-ray crystallography of 100 K is not optimal in terms of minimizing radiation damage and thereby increasing the structural information obtainable in a single experiment. At 50 K, specific radiation damage to disulfide bridges is reduced by a factor of 4 compared to 100 K, and samples can tolerate a factor of 2.6 and 3.9 higher dose, as judged by the increase of R free values of elastase and cubic insulin crystals, respectively.

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Cryoradiolytic reduction of crystalline heme proteins: analysis by UV-Vis spectroscopy and X-ray crystallography

Cited by 141 publications

References 75 publications

Temperature-dependent macromolecular X-ray crystallography

Temperature-dependent macromolecular X-ray crystallography

Crystal Structure and Biochemical Features of EfeB/YcdB from Escherichia coli O157

Origin and temperature dependence of radiation damage in biological samples at cryogenic temperatures

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