Abstract：
:Enhanced Green Fluorescent Protein (EGFP) is a variant of wild-type Green Fluorescent Protein from the jellyfish Aequorea victoria, whose mutations S65T and F64L increase brightness and folding efficiency. EGFP is extensively used in cell biology and biochemistry as a colocalization or expression reporter. Surprisingly, the structure of this very popular protein has not been determined yet. We report here its crystallographic structure at 1.5Å resolution which shows significant differences in the vicinity of residue 64 and of the chromophore. In particular, two conformations are observed for the key residue glutamic acid 222, in apparent contradiction with the single fluorescence lifetime of the protein. We then show that X-ray induced decarboxylation of Glu222 during diffraction data collection results in the disruption of a hydrogen-bond network near the chromophore. Using single-crystal microspectrophotometry, we demonstrate that this correlates with a significant loss of the fluorescence properties. We thus propose a mechanism of bleaching of the protein at low temperature. Taken together, these two sets of results highlight the stabilizing role of Glu222 to the chromophore cavity of EGFP.
journal_name
J Struct Bioljournal_title
Journal of structural biologyauthors
Royant A,Noirclerc-Savoye Mdoi
10.1016/j.jsb.2011.02.004subject
Has Abstractpub_date
2011-05-01 00:00:00pages
385-90issue
2eissn
1047-8477issn
1095-8657pii
S1047-8477(11)00043-8journal_volume
174pub_type
杂志文章abstract：:In order to successfully perform the 3D reconstruction in electron tomography, transmission electron microscope images must be accurately aligned or registered. So far, the problem is solved by either manually showing the corresponding fiducial markers from the set of images or automatically using simple correlation b...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1006/jsbi.2001.4343
更新日期：2001-01-01 00:00:00
abstract：:Collagen is the most important structural protein of the animal body. Its unique triple-helix structure and extremely high level of crystallinity make it exceptionally efficient in generating the second harmonic of incident light, and we show here how this leads to a novel mode of microscopy of immediate practical sig...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/s1047-8477(02)00576-2
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abstract：:The use of a Zernike-type phase plate in biologic cryo-electron microscopy allows the imaging, without using defocus, of what are predominantly phase objects. It is thought that such phase-plate implementations might result in higher quality images, free from the problems of CTF correction that occur when images must ...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2011.03.020
更新日期：2011-06-01 00:00:00
abstract：:A defining property of a crystal is its symmetry. This mini-review sets out to summarize all aspects that define 2D crystallographic symmetry as applied to the study of macromolecular structure. It begins by defining molecular point symmetries, before covering crystallographic symmetry operations in 2D, common notatio...
journal_title：Journal of structural biology
pub_type： 杂志文章,评审
doi：10.1016/j.jsb.2007.07.002
更新日期：2007-12-01 00:00:00
abstract：:The fungal pathogen Cryptococcus neoformans is a causative agent of meningoencephalitis in humans. For its pathogenicity, the inositol polyphosphate biosynthetic pathway plays critical roles. Recently, Ipk1 from C. neoformans (CnIpk1) was identified as an inositol 1,3,4,5,6-pentakisphosphate 2-kinase that catalyzes th...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2017.09.004
更新日期：2017-11-01 00:00:00
abstract：:The 32 kDa hydrophilic and acidic enamelin, the most stable cleavage fragment of the enamel specific glycoprotein, is believed to play vital roles in controlling crystal nucleation or growth during enamel biomineralization. Circular dichroism and Fourier transform infrared spectra demonstrate that the secondary struct...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2008.04.007
更新日期：2008-07-01 00:00:00
abstract：:Human coagulation factor VIIai that lacks the Gla domain (residues 1-44) has been prepared, purified, and crystallised. First, recombinant factor VII was activated to form factor VIIa, the active site was then inhibited with 1,5-dansyl-Glu-Gly-Arg-chloromethyl ketone, and finally the Gla domain was removed by chymotry...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1006/jsbi.1998.4078
更新日期：1999-03-01 00:00:00
abstract：:A new scanning probe microscope, the photonic force microscope (PFM), based on optical tweezers and two-photon absorption processes for biological applications is described. Optical tweezers are used to trap a fluorescent latex bead with a diameter of 200 nm in an aqueous solution in all three dimensions. The fluoresc...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1006/jsbi.1997.3880
更新日期：1997-07-01 00:00:00
abstract：:The emergence of electron tomography as a tool for three dimensional structure determination of cells and tissues has brought its own challenges for the preparation of thick sections. High pressure freezing in combination with freeze substitution provides the best method for obtaining the largest volume of well-preser...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2007.09.002
更新日期：2008-03-01 00:00:00
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journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2014.03.022
更新日期：2014-06-01 00:00:00
abstract：:The molecular mechanism underlining the antibacterial activity of the bacteriocin AS-48 is not known, and two different and opposite alternatives have been proposed. Available data suggested that the interaction of positively charged amino acids of AS-48 with the membrane would produce membrane destabilization and dis...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2015.03.006
更新日期：2015-05-01 00:00:00
abstract：:Single-axis cryo-electron tomography of vitrified specimens has become a method of choice to reconstruct in three dimensions macromolecular assemblies in their cellular context or prepared from purified components. Here, we asked how a dual-axis acquisition scheme would improve three-dimensional reconstructions of mic...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2012.11.004
更新日期：2013-02-01 00:00:00
abstract：:Upon illumination the visual receptor rhodopsin (Rho) transitions to the activated form Rho(∗), which binds the heterotrimeric G protein, transducin (Gt) causing GDP to GTP exchange and Gt dissociation. Using succinylated concanavalin A (sConA) as a probe, we visualized native Rho dimers solubilized in 1mM n-dodecyl-β...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2013.02.014
更新日期：2013-05-01 00:00:00
abstract：:We have developed a new two-step algorithm to determine the astigmatism of images from transmission electron microscopes (TEMs). Instead of computing the radial average of the power spectrum, we divide the power spectrum of a TEM image 1 to m (typically 32) sectors. We use a technique based on perturbation analysis of...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2006.07.021
更新日期：2007-01-01 00:00:00
abstract：:We present a sample preparation method for cryo-electron microscopy (cryo-EM) that requires only 3-20nL of sample to prepare a cryo-EM grid, depending on the protocol used. The sample is applied and spread on the grid by a microcapillary. The procedure does not involve any blotting steps, and real-time monitoring allo...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2016.11.002
更新日期：2017-03-01 00:00:00
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journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2011.10.011
更新日期：2012-02-01 00:00:00
abstract：:Structures of biological macromolecules determined by transmission cryoelectron microscopy (cryo-TEM) and three-dimensional image reconstruction are often displayed as surface-shaded representations with depth cueing along the viewed direction (Z cueing). Depth cueing to indicate distance from the center of virus part...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1006/jsbi.1997.3902
更新日期：1997-10-01 00:00:00
abstract：:Eosinophils, basophils, and mast cells produce and secrete active substances whose role is to attack invading parasites and protect the host. In this study we use morphometric methods to study mast cells in the blind mole rat (Spalax ehrenbergi). The subterranean and solitary way of life of this species has led to the...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1006/jsbi.2001.4429
更新日期：2001-11-01 00:00:00
abstract：:Tobacco streak virus (TSV), the type member of Ilarvirus genus, is a major plant pathogen. TSV purified from infected plants consists of a ss-RNA genome encapsidated in spheroidal particles with diameters of 27, 30 and 33nm constructed from multiple copies of a single species of coat protein (CP) subunits. Apart from ...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2015.12.007
更新日期：2016-02-01 00:00:00
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journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2016.03.001
更新日期：2016-06-01 00:00:00
abstract：:Single crystals of UDP-N-acetylglucosamine enolpyruvyltransferase of Enterobacter cloacae have been grown by vapor diffusion using phosphate buffer as the precipitant. The crystals belong to the monoclinic space group C2 with a = 86.9 A, b = 155.9 A, c = 83.8 A, beta = 91.6 degrees. Assuming two monomers per asymmetri...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1006/jsbi.1996.0071
更新日期：1996-07-01 00:00:00
abstract：:DNA regions close to the origin of replication were visualized by the green fluorescent protein (GFP)-Lac repressor/lac operator system. The number of oriC-GFP fluorescent spots per cell and per nucleoid in batch-cultured cells corresponded to the theoretical DNA replication pattern. A similar pattern was observed in ...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2005.06.004
更新日期：2005-09-01 00:00:00
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journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2014.09.008
更新日期：2014-11-01 00:00:00
abstract：:Radiation damage is the primary factor that limits resolution in electron cryo-microscopy (cryo-EM) of frozen-hydrated biological samples. Negative effects of radiation damage are attenuated by cooling specimens to cryogenic temperatures using liquid nitrogen or liquid helium. We have examined the relationship between...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2009.11.001
更新日期：2010-03-01 00:00:00
abstract：:Mononuclear macrophages are important immune cells in the organisms. The complicated membrane structure underlying the diverse functions of mononuclear-macrophage has been largely unresolved. As a representative of monocyte-derived macrophages, the membrane structure of PMA differentiated THP-1 cells was comprehensive...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2019.04.002
更新日期：2019-06-01 00:00:00
abstract：:We have developed a simplified, efficient approach for the 3D reconstruction and analysis of mammalian cells in toto by electron microscope tomography (ET), to provide quantitative information regarding 'global' cellular organization at approximately 15-20 nm resolution. Two insulin-secreting beta cells-deemed 'functi...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2007.09.015
更新日期：2008-03-01 00:00:00
abstract：:A novel type 1 ribosome-inactivating protein (RIP) designated cucurmosin was isolated from the sarcocarp of Cucurbita moschata (pumpkin). Besides rRNA N-glycosidase activity, cucurmosin exhibits strong cytotoxicities to three cancer cell lines of both human and murine origins, but low toxicity to normal cells. Plant g...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2008.06.011
更新日期：2008-10-01 00:00:00
abstract：:Chaperone-proteases are responsible for the processive breakdown of proteins in eukaryotic, archaeal and bacterial cells. They are composed of a cylinder-shaped protease lined on the interior with proteolytic sites and of ATPase rings that bind to the apical sides of the protease to control substrate entry. We present...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2009.07.003
更新日期：2009-11-01 00:00:00
abstract：:The massive basal skeleton of a few remnant living hypercalcified sponges rediscovered since the 1960s are valuable representatives of ancient calcium carbonate biomineralization mechanisms in basal Metazoa. A multi-scale mineralogical characterization of the easily accessible Mediterranean living hypercalcified spong...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1016/j.jsb.2011.08.008
更新日期：2011-12-01 00:00:00
abstract：:Gramicidin is a polypeptide antibiotic which forms dimeric channels specific for the transport of monovalent cations across membranes. It adopts several different conformations, most notably double helical (pore) and helical dimer (channels) forms, which have very different structural and functional characteristics. T...
journal_title：Journal of structural biology
pub_type： 杂志文章
doi：10.1006/jsbi.1997.3948
更新日期：1998-01-01 00:00:00
