Radiation damage of proteins in the solid state: Changes of amino acid composition in catalase

Baumeister, Wolfgang; Hahn, M.; Seredynski, J.; Herbertz, L

doi:10.1016/0304-3991(76)90052-8

Cited by 15 publications

(3 citation statements)

References 9 publications

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“…In the absence of appreciable heating, it shows no significant dose-rate dependence over the range of fluxes available at third-generation synchrotron sources. Examination of electron-density maps has revealed preferential damage at particular sites within the protein (Weik et al, 2000;Burmeister, 2000;Leiros et al, 2001;Schiltz et al, 2004), consistent with studies on proteins in solution (Dertinger & Jung, 1970;Box, 1972;Houee-Levin & Sicard-Roselli, 2001) and in the solid state (Baumeister et al, 1976;Garrison, 1987).…”

Section: Introductionmentioning

confidence: 53%

Quantifying X-ray radiation damage in protein crystals at cryogenic temperatures

Kmetko

Husseini

Naides

et al. 2006

Acta Crystallogr D Biol Cryst

149

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The dependence of radiation damage to protein crystals at cryogenic temperatures upon the X-ray absorption cross-section of the crystal has been examined. Lysozyme crystals containing varying heavy-atom concentrations were irradiated and diffraction patterns were recorded as a function of the total number of incident photons. An experimental protocol and a coefficient of sensitivity to absorbed dose, proportional to the change in relative isotropic B factor, are defined that together yield a sensitive and robust measure of damage. Radiation damage per incident photon increases linearly with the absorption coefficient of the crystal, but damage per absorbed photon is the same for all heavy-atom concentrations. Similar damage per absorbed photon is observed for crystals of three proteins with different molecular sizes and solvent contents.

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Section: Introductionmentioning

confidence: 53%

Quantifying X-ray radiation damage in protein crystals at cryogenic temperatures

Kmetko

Husseini

Naides

et al. 2006

Acta Crystallogr D Biol Cryst

149

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“…There remains one to two angstroms of in-plane motion during the first 4 e − /Å 2 on gold substrates. These first few electrons are critical as they potentially contain the most high-resolution information (26, 27). Future work will focus on substrate design and image acquisition conditions to reduce the initial motion even more.…”

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confidence: 99%

Ultrastable gold substrates for electron cryomicroscopy

Russo

Passmore

2014

Science

371

322

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Despite recent advances, the structures of many proteins cannot be determined by electron cryomicroscopy because the individual proteins move during irradiation. This blurs the images so they cannot be aligned with each other to calculate a 3D density. Much of this movement stems from instabilities in the carbon substrates used to support frozen samples in the microscope. Here we demonstrate a new gold specimen support that nearly eliminates substrate motion during irradiation. This increases the sub-nanometer image contrast such that α-helices of individual proteins are resolved. With this improvement we determine the structure of apoferritin, a smooth, octahedral shell of α-helical subunits that is particularly difficult to solve by electron microscopy.This advance in substrate design will enable the solution of currently intractable protein structures.Recent developments in electron cryomicroscopy (cryo-EM) have allowed structure determination to near-atomic resolution for some macromolecular complexes (1-3). Still, many small and challenging structures cannot be determined by current cryo-EM methods to the resolutions required for accurate modeling of atomic structure. This is because electron micrograph quality still falls short of the physical limits imposed by radiation damage to the macromolecules (4-6). Reduced image quality likely has two primary origins: Inefficient detection of imaging electrons (6, 7), and motion and blurring of the particles during irradiation (6,(8)(9)(10)(11). Recent developments in electron detectors have addressed the first constraint (12,13), and have enabled the development of motion correction algorithms to ameliorate the effects of the second (14-16). Direct electron detectors can acquire images using fractions of the electron dose previously required, thus allowing the measurement of single molecule positions in time. This allows accurate tracking of large ensembles of particles, and offers a way to determine the physical origins of radiation-induced particle movement (14-17).Conventional amorphous carbon (am-C) substrates undergo bending and deformation during irradiation (9,11), which includes movement both parallel and perpendicular to the substrate plane. Incorporation of titanium-silicon, doped silicon carbide or graphene into substrate designs reduced radiation-induced specimen motion (17)(18)(19). Still these designs did not stop substrate movement and are challenging to manufacture and use. Here, we demonstrate a new cryo-EM support that nearly eliminates the radiation-induced deformation of thin, iceembedded specimens at cryogenic temperatures. This curtails the perpendicular and in-plane * To whom correspondence should be addressed; passmore@mrc-lmb.cam.ac.uk. Europe PMC Funders Group Europe PMC Funders Author ManuscriptsEurope PMC Funders Author Manuscripts components of motion during imaging, and thus improves image quality for all radiationsensitive cryogenic specimens.The support comprises a regular array of micron sized holes in a ≃500 Å thick foil ...

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“…The rapid destruction of high resolution specimen detail by the electron beam itself was first recognized and demonstrated by Williams & Fisher (1970) and subsequently by numerous others (Glaeser, 1971(Glaeser, , 1975Baumeister et al, 1976). The destructive mechanisms have been studied (Stenn & Bahr, 1970) and although the susceptibility of different specimens varies, it has been shown that in general doses even as low as 50-100 electrons/nm2 destroy all detail finer than 1 nm (Unwin & Henderson, 1975).…”

Section: Introductionmentioning

confidence: 98%

Combining accurate defocus with low‐dose imaging in high resolution electron microscopy of biological material

Wrigley

Brown

Chillingworth

1983

Journal of Microscopy

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SUMMARY High resolution (< 2 nm) electron microscopy of biological specimens requires three exacting conditions to be met simultaneously: (a) fine specimen detail must be protected from destruction by the electron beam (low dose), (b) the electron optics must be adjusted to be capable of imaging that detail interpretably (accurate defocus), and (c) a suitable field of interest must be identified. We describe a method encompassing all three with an 80% success rate using only minor modifications to a transmission electron microscope, and no expensive on‐line computing.

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Radiation damage of proteins in the solid state: Changes of amino acid composition in catalase

Cited by 15 publications

References 9 publications

Quantifying X-ray radiation damage in protein crystals at cryogenic temperatures

Quantifying X-ray radiation damage in protein crystals at cryogenic temperatures

Ultrastable gold substrates for electron cryomicroscopy

Combining accurate defocus with low‐dose imaging in high resolution electron microscopy of biological material

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