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Annular Pore
(18 References)
Lashuel, H. A., D. M. Hartley, et al. (2003). "Mixtures of wild-type and a pathogenic (E22G) form of Abeta40 in vitro accumulate protofibrils, including amyloid pores." J Mol Biol 332(4): 795-808.
Although APP mutations associated with inherited forms of Alzheimer's disease (AD) are relatively rare, detailed studies of these mutations may prove critical for gaining important insights into the mechanism(s) and etiology of AD. Here, we present a detailed biophysical characterization of the structural properties of protofibrils formed by the Arctic variant (E22G) of amyloid-beta protein (Abeta40(ARC)) as well as the effect of Abeta40(WT) on the distribution of the protofibrillar species formed by Abeta40(ARC) by characterizing biologically relevant mixtures of both proteins that may mimic the situation in the heterozygous patients. These studies revealed that the Arctic mutation accelerates both Abeta oligomerization and fibrillogenesis in vitro. In addition, Abeta40(ARC) was observed to affect both the morphology and the size distribution of Abeta protofibrils. Electron microscopy examination of the protofibrils formed by Abeta40(ARC) revealed several morphologies, including: (1) relatively compact spherical particles roughly 4-5 nm in diameter; (2) annular pore-like protofibrils; (3) large spherical particles 18-25 nm in diameter; and (4) short filaments with chain-like morphology. Conversion of Abeta40(ARC) protofibrils to fibrils occurred more rapidly than protofibrils formed in mixed solutions of Abeta40(WT)/Abeta40(ARC), suggesting that co-incubation of Abeta40(ARC) with Abeta40(WT) leads to kinetic stabilization of Abeta40(ARC) protofibrils. An increase in the ratio of Abeta(WT)/Abeta(MUT(Arctic)), therefore, may result in the accumulation of potential neurotoxic protofibrils and acceleration of disease progression in familial Alzheimer's disease mutation carriers.
Hinshaw, J. E. and R. A. Milligan (2003). "Nuclear pore complexes exceeding eightfold rotational symmetry." J Struct Biol 141(3): 259-68.
Nuclear pore complexes are rotationally symmetric structures that span the nuclear envelope and provide channels for nucleocytoplasmic traffic. These large complexes normally consist of eight spokes arranged around a central channel, although, occasionally, 9- and 10-fold nuclear pore complexes are found in preparations of Xenopus oocyte macronuclei. Here we examine these unusual nuclear pore complexes by negative stain electron microscopy and image analysis and compare the results with data previously obtained from 8-fold structures. The details in two-dimensional and three-dimensional maps indicate that the substructure of the spoke is the same in 8-, 9- and 10-fold nuclear pore complexes: therefore, the spoke is likely an immutable structural component. In all three variant forms, the spacing between adjacent annular subunits, which surround the central channel, is identical. Distances between spokes at higher radius decrease in the 9- and 10-fold nuclear pore complexes. These data imply that the most important connections holding the nuclear pore complex together are those between adjacent annular subunits and that these interactions may play a predominant role in nuclear pore complex assembly. Circumferential connections mediated by ring subunits and radial arms presumably further stabilize the structure and are flexible enough to accommodate additional spokes.
Shi, C. and N. M. Soldatov (2002). "Molecular determinants of voltage-dependent slow inactivation of the Ca2+ channel." J Biol Chem 277(9): 6813-21.
Ba(2+) current through the L-type Ca(2+) channel inactivates essentially by voltage-dependent mechanisms with fast and slow kinetics. Here we found that slow inactivation is mediated by an annular determinant composed of hydrophobic amino acids located near the cytoplasmic ends of transmembrane segments S6 of each repeat of the alpha(1C) subunit. We have determined the molecular requirements that completely obstruct slow inactivation. Critical interventions include simultaneous substitution of A752T in IIS6, V1165T in IIIS6, and I1475T in IVS6, each preventing in additive manner a considerable fraction of Ba(2+) current from inactivation. In addition, it requires the S405I mutation in segment IS6. The fractional inhibition of slow inactivation in tested mutants caused an acceleration of fast inactivation, suggesting that fast and slow inactivation mechanisms are linked. The channel lacking slow inactivation showed approximately 45% of the sustained Ba(2+) or Ca(2+) current with no indication of decay. The remaining fraction of the current was inactivated with a single-exponential decay (pi(f) approximately 10 ms), completely recovered from inactivation within 100 ms and did not exhibit Ca(2+)-dependent inactivation properties. No voltage-dependent characteristics were significantly changed, consistent with the C-type inactivation model suggesting constriction of the pore as the main mechanism possibly targeted by Ca(2+) sensors of inactivation.
Lashuel, H. A., D. Hartley, et al. (2002). "Neurodegenerative disease: amyloid pores from pathogenic mutations." Nature 418(6895): 291.
Alzheimer's and Parkinson's diseases are associated with the formation in the brain of amyloid fibrils from beta-amyloid and alpha-synuclein proteins, respectively. It is likely that oligomeric fibrillization intermediates (protofibrils), rather than the fibrils themselves, are pathogenic, but the mechanism by which they cause neuronal death remains a mystery. We show here that mutant amyloid proteins associated with familial Alzheimer's and Parkinson's diseases form morphologically indistinguishable annular protofibrils that resemble a class of pore-forming bacterial toxins, suggesting that inappropriate membrane permeabilization might be the cause of cell dysfunction and even cell death in amyloid diseases.
Lashuel, H. A., B. M. Petre, et al. (2002). "Alpha-synuclein, especially the Parkinson's disease-associated mutants, forms pore-like annular and tubular protofibrils." J Mol Biol 322(5): 1089-102.
Two mutations in the alpha-synuclein gene (A30P and A53T) have been linked to autosomal dominant early-onset Parkinson's disease (PD). Both mutations promote the formation of transient protofibrils (prefibrillar oligomers), suggesting that protofibrils are linked to cytotoxicity. In this work, the effect of these mutations on the structure of alpha-synuclein oligomers was investigated using electron microscopy and digital image processing. The PD-linked mutations (A30P and A53T) were observed to affect both the morphology and the size distribution of alpha-synuclein protofibrils (measured by analytical ultracentrifugation and scanning transmission electron microscopy). The A30P variant was observed to promote the formation of annular, pore-like protofibrils, whereas A53T promotes formation of annular and tubular protofibrillar structures. Wild-type alpha-synuclein also formed annular protofibrils, but only after extended incubation. The formation of pore-like oligomeric structures may explain the membrane permeabilization activity of alpha-synuclein protofibrils. These structures may contribute to the pathogenesis of PD.
Ding, T. T., S. J. Lee, et al. (2002). "Annular alpha-synuclein protofibrils are produced when spherical protofibrils are incubated in solution or bound to brain-derived membranes." Biochemistry 41(32): 10209-17.
The Parkinson's disease substantia nigra is characterized by the loss of dopaminergic neurons and the presence of cytoplasmic fibrillar Lewy bodies in surviving neurons. The major fibrillar protein of Lewy bodies is alpha-synuclein. Two point mutations in the alpha-synuclein gene are associated with autosomal-dominant Parkinson's disease (FPD). Studies of the in vitro fibrillization behavior of the mutant proteins suggest that fibril precursors, or alpha-synuclein protofibrils, rather than the fibrils, may be pathogenic. Atomic force microscopy (AFM) revealed two distinct forms of protofibrillar alpha-synuclein: rapidly formed spherical protofibrils and annular protofibrils, which were produced on prolonged incubation of spheres. The spherical protofibrils bound to brain-derived membrane fractions much more tightly than did monomeric or fibrillar alpha-synuclein, and membrane-associated annular protofibrils were observed. The structural features of alpha-synuclein annular protofibrils are reminiscent of bacterial pore-forming toxins and are consistent with their porelike activity in vitro. Thus, abnormal membrane permeabilization may be a pathogenic mechanism in PD.
Tadjibaeva, G., R. Sabirov, et al. (2000). "Flammutoxin, a cytolysin from the edible mushroom Flammulina velutipes, forms two different types of voltage-gated channels in lipid bilayer membranes." Biochim Biophys Acta 1467(2): 431-43.
Flammutoxin, a 31-kDa cardiotoxic and cytolytic protein from the edible mushroom Flammulina velutipes, has been shown to assemble into a pore-forming annular oligomer with outer and inner diameters of 10 and 5 nm on the target cells [Tomita et al., Biochem. J. 333 (1998) 129-137]. Here we studied electrophysiological properties of flammutoxin channels using planar lipid bilayer technique, and found that flammutoxin formed two types of moderately cation-selective, voltage-gated channels with smaller and larger current amplitudes (1-4.5 pA and 20-30 pA, respectively, at 20 mV) in the lipid bilayers composed of phospholipid and cholesterol. The larger-conductance single channel showed the properties of a wide water-filled pore such as a linear relationship between channel conductance and salt concentration of the bathing solution. The functional diameter of the larger-conductance channel was estimated to be 4-5 nm by measuring the current conductance in the presence of polyethylene glycols of various sizes. In contrast, the smaller-conductance single channels showed a non-linear current to voltage curve and a saturating conductance to increasing salt concentration. These results suggest that the larger-conductance channel of flammutoxin corresponds to the hemolytic pore complex, while the smaller-conductance channel may reflect the intermediate state(s) of the assembling toxin.
Ortells, M. O. and G. G. Lunt (1996). "A mixed helix-beta-sheet model of the transmembrane region of the nicotinic acetylcholine receptor." Protein Eng 9(1): 51-9.
We have modelled the transmembrane region of the alpha 7 nicotinic acetylcholine receptor as a mixed alpha-helical/beta-sheet structure. The model was mainly based on the crystal structure of a pore-forming toxin, heat-labile enterotoxin. This is a pentameric protein having a central pore or channel composed of five alpha-helices, one from each of the 5 B subunits that form this pentamer. The remainder of this structure is beta-sheet, loops and a short alpha-helix, not included in the model. The model uses this channel as a template to build the transmembrane region, from M1 to the middle of M3. The remainder of M3 and M4 were built de novo as alpha-helices. Great consideration was given to labelling data available for the transmembrane region. In general terms, the shape of the model agrees very well with that obtained independently by electron microscopic analysis and the secondary structure predicted by the model is in accord with that estimated independently by Fourier transform infrared spectroscopy. The M2 helical region of the model is only slightly kinked, contrary to what is inferred from electron microscopic analysis, but has the same overall shape and form. On the membrane face of the model, the presence of deep pockets may provide the structural basis for the distinction between annular and non-annular lipid binding sites. Also, the transmembrane region is clearly asymmetric in the direction perpendicular to the membrane, and this may have strong influence on the surrounding lipid composition of each leaflet of the cytoplasmic membrane.
Fujimoto, K. and P. Pinto da Silva (1989). "Surface views of nuclear pores in isolated rat liver nuclei as revealed by fracture-flip/Triton-X." Eur J Cell Biol 50(2): 390-7.
We report here the use of fracture-flip (Anderson Forsman, C., P. Pinto da Silva, J. Cell Sci. 90, 531-541 (1988] and fracture-flip/Triton X-100 to examine the cytoplasmic surface of the outer nuclear membrane, and the perinuclear (exoplasmic) surface of the inner nuclear membrane of rat liver nuclei. After freeze-fracture, carbon stabilization and thawing, unfixed nuclei were treated with Triton X-100 to dissolve unfractured portions of the outer nuclear membrane. Flipping of the casts with their carbon-stabilized membrane halves (cytoplasmic half of the outer nuclear membrane; exoplasmic half of the inner nuclear membrane) exposed the cytoplasmic surface of the outer nuclear membrane and the surface of the inner nuclear membrane facing the perinuclear space. After drying, these membrane surfaces were visualized by Pt/C shadowing. We show that the cytoplasmic surface of outer nuclear membrane is covered by numerous large, oblong "kernel-like" globular particles (30-35 nm length, 25-30 nm width), often aligned into curved chains or grouped into clusters. Because of their size and characteristic arrays, we interpret these particles as ribosomes. Over the cytoplasmic surfaces of the outer nuclear membrane the nuclear pore complexes (outer diameter: 120-130 nm) appear as relatively inconspicuous annular structures, only slightly raised against the surrounding areas of membrane surface. Observation of the surface of the inner nuclear membrane facing the perinuclear space reveals prominent, sharply raised, "doughnut-like" nuclear pore complexes (outer diameter: 120-130 nm; inner diameter: 40 nm). Our results indicate the absence of large, protruding, ribosome-like structures lining the nuclear pore at its cytoplasmic surface, as proposed before.(ABSTRACT TRUNCATED AT 250 WORDS)
Olofsson, A., U. Kaveus, et al. (1988). "The projection structure of alpha-toxin from Staphylococcus aureus in human platelet membranes as analyzed by electron microscopy and image processing." J Ultrastruct Mol Struct Res 100(2): 194-200.
Most strains of Staphylococcus aureus produce alpha-toxin, a 33-kDa membrane active protein which is considered to be an important virulence factor of this bacterium. When alpha-toxin interacts with membranes an oligomeric from of the toxin can be seen by electron microscopy as characteristic ring structures in the membrane. A two-dimensional study of these annular structures, incorporated in membranes of human platelets, was performed, introducing a partly new method for rotational alignment of individual particles. It is shown that the averaged oligomer consists of six subunits. At neutral pH the outer diameter of the ring is about 75 A. The stain-filled pore or cavity in the center has a diameter of about 25 A. The size of the hexamer is increased if the pH is lowered.
Troitskaia, L. P., S. N. Kuz'mina, et al. (1981). "[Electron microscope characteristics of the nuclear matrix and its fractions]." Tsitologiia 23(6): 620-5.
The rat liver nuclear matrix retains the shape of the nucleus and reveals a sponge-like structure in negative staining and scanning electron microscopy. A fibrous layer (dense lamina) with associated pore complexes are preserved on the surface of the nuclear matrix. The cytoplasmic face of the nuclear matrix is perceived as a network consisting of cells (or units) of 10-30 nm in diameter in negative staining as well as in high resolution scanning electron microscopy. In sections, a fibrous layer, 15-30 nm in width with granules of 7-10 nm in diameter, can be observed. In pore complexes associated with the fibrous layer granular and fibrillar components rather than central granules are observed. The pore complexes differ in arrangement of the annular granules. Structures similar to pore complexes are revealed in close proximity to the nucleoli. The biogenesis of the pore complexes is discussed. A few morphologically different structures could be derived be fractionation of the nuclear matrix. A fraction rich in pore complexes, and a fraction retaining the shape of the nucleus with spongy or alveolar structure were isolated. The latter fraction is regarded to form a protein framework or skeleton of the nucleus.
Kuzmina, S., T. Buldyaeva, et al. (1981). "Characterization and fractionation of rat liver nuclear matrix." Eur J Cell Biol 25(2): 225-32.
The rat liver nuclear matrix retains the shape of the nucleus and reveals a sponge-like structure in negative staining and scanning electron microscopy. A fibrous layer (dense lamina) with associated pore complexes is preserved on the surface of the nuclear matrix. In negative staining as well as in high resolution scanning electron microscopy this layer is perceived as a network consisting of alveoli of 10 to 30 nm in diameter with pore complexes differing in arrangement of the annular granules. In sections a fibrous layer 15 to 30 nm in width granules of 7 to 10 nm in diameter can be observed. Structures similar to the pore complexes are revealed not only at the fibrous layer but also inside the nucleus and in close proximity to the nucleoli. Four fractions of the nuclear matrix have been isolated by successive extraction with 0.05 M EDTA and 0.025 N NaOH. Almost 80% of the total matrix protein dissolves in dilute alkali. 4 to 5% of the total matrix protein is soluble in EDTA. From insoluble residue two fractions can be isolated: one rich in pore complexes and another one retaining the shape of the nucleus with spongy or alveolar structure. The latter fraction is regarded to constitute a proper framework or skeleton of the nucleus.
Vorbrodt, A. and G. G. Maul (1980). "Cytochemical studies on the relation of nucleoside triphosphatase activity to ribonucleoproteins in isolated rat liver nuclei." J Histochem Cytochem 28(1): 27-35.
Cytochemical tests for nucleosidetriphosphatase (NTPase) and Bernhard's preferential staining for ribonucleoproteins (RNP) were applied to isolated rat liver nuclei. The strongest and most easily reproducible positive reaction for NTPase was detected at pH 7.7 with ATP and GTP. This reaction was activated by Mg2+ and Ca2+ and inhibited by Be2+, Zn2+, quercetin, and ribonuclease. The major sites of enzyme reaction were intranuclear RNA-containing structures. Incubation of nuclei in ATP-stimulated RNA-release medium eliminated a considerable part of the material showing both NTPase reaction and staining for RNP; the perichromatin granules disappeared, while interchromatin granules remained. NTPase activity in the nuclear envelope seems to be associated with the annular part of nuclear pore complexes (permanent component) and with RNP particles translocated through nuclear pores or attached to the surface of nuclei (transitional component). From a morphological point of view, these observations support previous biochemical data suggesting the existence of a connection between NTPase activity and the translocation of RNP particles through the nuclear envelope.
Chelidze, P. V., G. D. Tumanishvili, et al. (1980). "[Effect of low temperature on the ultrastructure of hepatocyte nuclei in 11-day-old chick embryos]." Tsitologiia 22(10): 1146-52.
A study was made of the ultrastructure of nuclei of 11-day old chick embryos exposed to low temperature (-15 degrees C) for various periods of time and returned to the normal conditions of incubations (+37 degrees). The exposure to the low temperature induced some drastic ultrastructural alterations in the nuclear pore complex that involved the loss of lateral components of annulus (annular granules) and of the central granule. Some pores displayed a diaphragm or its structures (peripheral granules), whereas in other pores the pore complex material was seen totally disorganized. The degree of chromatin condensation in such nuclei increased. After the normal condition of chick embryo incubation was resumed, the ultrastructural organization of the chromatin and the pore complex appeared to be normalized. The pore complex reorganization occurred one hour after heating. It is suggested that the abnormal pore structure may correspond to an inactive state of the pore.
Parfenov, V. N. (1979). "The karyosphere during late oogenesis in Rana ridibunda." Eur J Cell Biol 19(2): 102-8.
The organization of the nucleus in the oocytes of Rana ridibunda was examined during late diplotene at the light and electron microscopic level. At this stage the chromosomes are relatively condensed and assembled in the centre of the nucleus, constituting a karyosphere. The chromosomes here are associated with the central "protein sphere" (15--20 microns in diameter), obviously at their telomeres. Numerous nucleoli are accumulated around the chromosomes, forming a karyosphere capsule and contain segregated fibrillar and granular components; structures resembling perinucleolar chromatin and fibrillar bodies (spherules) are associated with the nucleoli. Granules 30 to 40 nm in diameter are seen to surround the fibrillar spherules. "Nucleolus-like bodies" consisting of granules 10 to 15 nm in diameter which are embedded in finely fibrillar material are often associated in contact with the chromosomes. The central sphere is an accumulation of annular structures similar to those of the pore complexes of the nuclear envelope. These structures are bound to the chromosome material, the "nucleolus-like bodies" and the fibrillar bodies. A participation of "nucleolus-like bodies" in the formation of the central sphere is suggested. A possible role of the nuclear protein matrix in the construction of the karyosphere elements is discussed.
Riley, D. E. and J. M. Keller (1978). "The ultrastructure of non-membranous nuclear ghosts." J Cell Sci 32: 249-68.
Non-membranous HeLa cell nuclear ghosts, representing non-membranous nuclear envelope or 'skeletal' components, have been examined in whole-mount fashion by transmission electron microscopy. Major components of the ghosts include annuli with inner and outer diameters of 43 and 90 nm, respectively, which are consistent in dimensions with nuclear pore complexes. Also present are rod-like images (260 nm in length and 50 nm in width or diameter) representing either previously unobserved nuclear structures, or condensations of repeating functional units not otherwise observable. The annular and rod-like images were also observed when various steps in the ghost isolation procedure, such as the use of detergents, 0.5 M MgCl2 and polylysine attachment of the ghosts to electron-microscope grids, were circumvented. The annular and rod-like images are connected into linear and polygonal arrays by strands (15-30 nm in width) that are sensitive to DNase I and DNase II but resistant to nuclease S1. Thus, although the non-membranous ghosts from HeLa cells are composed primarily of protein, enzymic dissection indicates that their gross integrity is markedly dependent on double-stranded DNA. Nuclear ghosts prepared from a wide range of species including mammals, birds and plants, exhibited essentially the same components and organization.
Goldstein, M. A. (1975). "Ultrastructural study of nuclear pores in hypertrophied heart cells." Recent Adv Stud Cardiac Struct Metab 6: 335-41.
Nuclear pores were consistently found in both spontaneous and experimentally induced hypertrophy. The structure of nuclear pores in hypertrophied hamster, guinea pig, and rabbit heart cells was similar. The morphology of nuclear pores was similar to that previously seen in normal cardiac cells and in other cell types. Each pore complex was composed of a pore sometimes containing a central granule, a pore margin having an octagonal shape, and two rings of annular granules lying upon the pore margin.
Hoeijmakers, J. H., J. H. Schel, et al. (1974). "Structure of the nuclear pore complex in mammalian cells. Two annular components." Exp Cell Res 87(1): 195-206.