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U Aebi

Publications and source records attributed to U Aebi.

At least 109 records · Page 6Linked to original sources
PubMed1995

Evidence of a defined spatial arrangement of hyaluronate in the central filament of cartilage proteoglycan aggregates.

Aggregates of proteoglycans from the Swarm rat chondrosarcoma reassembled in vitro have been studied by rotary-shadowing electron microscopy, and shown to be similar to native structures that have never been dissociated [Mörgelin, Engel, Heinegård and Paulsson (1992) J. Biol. Chem. 267, 14275-14284]. A hyaluronate with defined chain length (HAshort) has now been prepared by autoclaving high-Mr hyaluronate and fractionation to a narrow size distribution by gel filtration. Proteoglycan monomers, core protein, hyaluronate-binding region and link protein were combined with HAshort. Free chains of HAshort and reconstituted complexes with proteoglycan, link protein and aggrecan fragments were examined by electron microscopy after rotary shadowing. Length measurements showed that the hyaluronate was condensed to about half of its original length on binding intact aggrecan monomers, any aggrecan fragment or link protein alone. This strongly implies that hyaluronate adopts a defined spatial arrangement within the central filament of the aggregate, probably different from its secondary structure in solution. No differences in length were observed between link-free and link-stabilized aggregates.

Animals↗
PubMed1995

Ultrastructural modification of human skeletal muscle tissue with 6-month moderate-intensity exercise training.

The study was aimed at investigating if endurance training of moderate intensity and longer duration, intended to promote health rather than performance, evokes ultrastructural changes in skeletal muscle tissue comparable to those observed after high-intensity protocols. Twenty healthy, middle-aged men enrolled in a 6-month, home-based jogging program of 120 min/wk at 75% VO2max. Only 12 men showed a sufficient exercise adherence over the 6 months (> or = 60 min/wk on average) and were included into statistical analysis. Their average training activity was 105 +/- 31 min/wk. The results revealed significant increases in VO2max (+8.4%, p < 0.01) and submaximal power output (+18.1%, p < 0.01). Total mitochondrial volume density in M. vastus lateralis increased by 20% (p < 0.05) with a larger increase in subsarcolemmal volume compared to central volume (50% vs 15%). No changes in volume of intracellular lipid droplets, capillary density, capillary per fiber ratio, fiber mean cross-sectional area and muscle fiber type could be observed. Body composition analysis showed a decrease in trunk fat mass (-7.3%, p < 0.05) and an increase in trunk lean mass (+1.7%, p < 0.05), while changes in the legs were not significant. It can be concluded that a moderate-intensity, health oriented endurance training beneficially affected cardiovascular and muscle oxidation capacity as well as body composition in the trunk area. No adaptations in capillaries or lipid metabolism could be demonstrated. The results support the hypothesis of thresholds for induction of adaptation processes in muscle skeletal tissue depending on the intensity of the exercise stimulus.

Adipose Tissue↗
PubMed1995

Structural analysis of the p62 complex, an assembly of O-linked glycoproteins that localizes near the central gated channel of the nuclear pore complex.

The p62 complex is an oligomeric assembly of O-linked glycoproteins of the nuclear pore complex that interacts with cytosolic transport factors and is part of the machinery for nuclear protein import. In this study we have purified the p62 complex from rat liver nuclear envelopes and analyzed its structure and composition. The p62 complex consists of four distinct polypeptides (p62, p58, p54, and p45) and has a mass of approximately 234 kDa, calculated from its hydrodynamic properties and supported by chemical cross-linking and scanning transmission electron microscopy. These data suggest that the p62 complex contains one copy of each constituent polypeptide. Analysis of preparations of the p62 complex by electron microscopy using rotary metal shadowing and negative staining revealed donut-shaped particles with a diameter of approximately 15 nm. Immunogold electron microscopy of isolated rat liver nuclear envelopes demonstrated that p62 occurs on both the nucleoplasmic and cytoplasmic sides of the pore complex near the central gated channel involved in active transport of proteins and RNAs. The properties and localization of the p62 complex suggest that it may be involved in binding transport ligands near the center of the nuclear pore complex and in subsequently transferring them to the gated transport channel.

Animals↗
PubMed1995

Exploring nuclear pore complex structure and function in molecular detail.

Bidirectional molecular trafficking between the nucleus and the cytoplasm of eukaryotic cells occurs through the nuclear pore complexes (NPCs), approximately 120 megadalton supramolecular assemblies embedded in the double-membraned nuclear envelope. Significant progress has been made in elucidating the three-dimensional (3-D) architecture of the NPC, and in identifying, characterizing, and cloning and sequencing NPC proteins. Several of these have now been localized within the 3-D structure of the NPC. Nevertheless, there still remain major questions relating to the conformation, molecular composition and functional roles of distinct NPC components. Here we review recent structural studies from our group and others which have contributed toward dissecting the molecular architecture of the NPC. We also present our results on the molecular characterization of some NPC components, and on the elucidation of their functional roles in mediated nucleocytoplasmic transport.

Animals↗
PubMed1994

Towards atomic interpretation of F-actin filament three-dimensional reconstructions.

We have recorded dark field images of negatively stained F-actin filaments polymerized with 2 mM MgCl2 and 50 mM KCl with a scanning transmission electron microscope and computed 3-D reconstructions using a helical parameter search to optimize simultaneously the helical repeat length, the radial position of the filament axis, and the helical selection rule. The resulting optimized averaged filament 3-D reconstruction at 2.5 nm resolution is remarkably similar to an atomic model of the F-actin filament. By comparison, several structural features of the reconstruction can be interpreted at the level of distinct secondary structure elements, and predictions made by the atomic model could be verified: for instance, the density connecting the two long-pitch helical strands in our reconstruction co-localizes with an extended beta-hairpin, the "hydrophobic loop" (i.e. residues 262 to 274), which according to the atomic model establishes the major intersubunit contact between the two long-pitch helical strands. The most pronounced structural variations among individual filament 3-D reconstructions were observed in (1) the details of the intersubunit contact pattern between the two long-pitch helical strands, and (2) the exact size and shape of subdomain 2 of the F-actin molecule, which appears rather flexible and easily deformed. In addition, we found that all phenotypes of F-actin filament 3-D reconstructions that arise from small deviations from the optimal helical parameters or from lowering the nominal resolution exhibited stronger intersubunit contacts between than along the two long-pitch helical strands, a structural feature that has been emphasized for a number of F-actin filament 3-D reconstructions in the past. Since this is clearly at variance with the relative strength of the intersubunit contacts as predicted by the atomic model, it may represent an artifactual structural feature arising from low-resolution data or suboptimal helical data processing, and should therefore be interpreted with caution in terms of indicating chemical, mechanical or conformational states of the F-actin filament.

Actins↗
PubMed1994

Fibritin encoded by bacteriophage T4 gene wac has a parallel triple-stranded alpha-helical coiled-coil structure.

The bacteriophage T4 late gene wac (whisker's antigen control) encodes a fibrous protein which forms a collar/whiskers complex. Whiskers function as a helper protein for the long tail fibres assembly and plays a role in regulating retraction of the long tail fibres in response to environmental conditions. In this work we show that expression of the cloned wac gene in Escherichia coli yields a protein oligomer of 53 nm length which we call fibritin, and which is able to complement gpwac T4 particles in vitro. CD spectroscopy of fibritin indicates a 90% alpha-helical content, and scanning calorimetry shows that the protein has several distinct domains. The analysis of the 486 amino acid sequence of fibritin reveals three structural components: a 408 amino acid region that contains 12 putative coiled-coil segments with a canonical heptad (a-b-c-d-e-f-g)n substructure where the "a" and "d" positions are preferentially occupied by apolar residues, and the N and C-terminal domains (47 and 29 amino acid residues, respectively) have no heptad substructure. The distribution of hydrophobic residues within heptads is more similar to a triple than to a double coiled-coil. The alpha-helical segments are separated by short "linker" regions, variable in length, that have a high proportion of glycine and proline residues. Each coiled-coil segment has, on the borders with linker regions, residues that are common to the N and C-terminal caps of the alpha-helices. Full-length and amino-terminally truncated fibritins can be reassembled in vitro after temperature-induced denaturation. Co-assembly of full-length fibritin and the N-terminal deletion mutant, as well as analytical centrifugation, indicates that the protein is a parallel triple-standard alpha-helical coiled-coil. Deletions of various N-terminal portions of fibritin did not block trimerisation but the mutant trimers are unable to bind to T4 particles. The last 18 C-terminal residues of fibritin are required for correct trimerisation of gpwac monomers in vivo. We propose that fibritin might serve as a convenient model for the investigation of folding and assembly mechanisms of alpha-fibrous proteins.

Amino Acid Sequence↗
PubMed1994

Mammalian uroplakins. A group of highly conserved urothelial differentiation-related membrane proteins.

The asymmetric unit membrane (AUM) forms the apical plaques of mammalian urothelium and is believed to play a role in strengthening the urothelial apical surface thus preventing the cells from rupturing during bladder distention. We have shown previously that purified bovine AUMs contain four major integral membrane proteins: the uroplakins Ia (27 kDa), Ib (28 kDa), II (15 kDa), and III (47 kDa). This contradicts some previous reports indicating that some of these proteins are absent in AUMs of several species. Using an improved procedure, we isolated AUMs from, in addition to cattle, eight mammalian species (human, monkey, sheep, pig, dog, rabbit, rat, and mouse). The AUMs of these species appear morphologically similar bearing crystalline patches of 12-nm protein particles with a center-to-center spacing of 16.5 nm. Using antibodies raised against synthetic oligopeptides or individual bovine uroplakins, we established by immunoblotting that the four uroplakins are present in AUMs of all these species. The DNA-deduced amino acid sequences of bovine and mouse uroplakin II revealed 83% identity. These results indicate that uroplakins Ia, Ib, II, and III are the major protein components of probably all mammalian urothelial plaques, and that the sequence and three-dimensional structure of uroplakin molecules are highly conserved during mammalian evolution.

Amino Acid Sequence↗
PubMed1994

Making heads and tails of intermediate filament assembly, dynamics and networks.

Thus far, intermediate filaments (IFs) have been the least understood of the three cytoskeletal filament systems with regard to their structure, assembly, network formation, and dynamics. This picture is now slowly but definitely changing, as recent in vivo and in vitro experiments, including generation of transgenic animals, have yielded important new data shedding light on the following areas: the molecular architecture of IFs; the role of the highly variable end domains during IF assembly and network formation; the factors that govern whether IF proteins are involved in de novo filament formation or are incorporated into a pre-existing IF network; and the effects of post-translational modifications, such as phosphorylation and glycosylation of IF polypeptides, on filament assembly, dynamics and turnover.

Animals↗
PubMed1994

Interactions and three-dimensional localization of a group of nuclear pore complex proteins.

We have used antibodies directed against a number of nuclear pore complex (NPC) proteins to determine their mutual interactions and location within the three-dimensional structure of the NPC. A monoclonal antibody, termed QE5, recognized three NPC polypeptides, p250, NUP153, and p62 on Western blots, and labeled the nuclear envelope of several cultured cell lines by immunofluorescence microscopy. These three polypeptides contained O-linked N-acetylglucosamine residues and were released from the NPC by detergent/high-salt treatment as discrete high molecular weight complexes. p250 was found in association with a novel 75 kD protein, NUP153 was released as a homo-oligomer of about 1 megadalton, and p62 was associated with polypeptides of 58 and 54 kD (previously reported by Finlay, D. R., E. Meier, P. Bradley, J. Horecka, and D. J. Forbes. 1991. J. Cell Biol. 114:169-183). p75, p58, and p54 were not galactosylated in vitro. Xenopus oocyte NEs were labeled with gold-conjugated QE5 and prepared for electron microscopy by quick freezing/freeze drying/rotary metal shadowing. This EM preparation method enabled us to more precisely localize the epitopes of this antibody to the cytoplasmic filaments and the nuclear basket of the NPC. Since QE5 recognizes three O-linked NPC glycoproteins, its labeling was compared with that of the lectin wheat germ agglutinin which recognizes O-linked N-acetylglucosamine moieties. The two probes were found to yield similar, although not identical, distributions of label. To identify the individual proteins with particular NPC components, we have used an anti-peptide antibody against NUP153 and a monospecific anti-p250 polyclonal antibody. Labeling with these two antibodies has documented that NUP153 is a constituent of the nuclear basket with at least one of its epitopes residing in its terminal ring, whereas p250 is a constituent of the cytoplasmic filaments.

Amino Acid Sequence↗
PubMed1994

Tpr, a large coiled coil protein whose amino terminus is involved in activation of oncogenic kinases, is localized to the cytoplasmic surface of the nuclear pore complex.

From a panel of monoclonal antibodies raised against fractions of rat liver nuclear envelopes (NEs), we have identified an antibody, RL30, which reacts with novel nuclear pore complex (NPC) antigens that are not O-glycosylated. By immunofluorescence staining of cultured cells, RL30 reacts exclusively with the NE in a punctate pattern that largely coincides with that of identified NPC proteins. RL30 labels only the cytoplasmic surface of the NPC in immunogold electron microscopy, predominantly in peripheral regions nearby the cytoplasmic ring. In immunoblots of isolated rat liver NEs and cultured rat cells, RL30 recognizes a 265-kD band, as well as a series of 175-265-kD bands in rat liver NEs that are likely to be proteolytic products of p265. Sequencing of peptides from the 175- and 265-kD RL30 antigens of rat liver revealed that they are both closely related to human Tpr, a protein whose amino-terminal 150-250 amino acids appear in oncogenic fusions with the kinase domains of the met, trk, and raf protooncogenes. We found that in vitro translation of human Tpr mRNA yields a major 265-kD band. Considered together, these data indicate that the 265-kD RL30 antigen in the NPC is the rat homologue of Tpr. Interestingly, Tpr contains an exceptionally long predicted coiled coil domain (approximately 1600 amino acids). The localization and predicted structure of Tpr suggest that it is a component of the cytoplasmic fibrils of the NPC implicated in nuclear protein import. Immunofluorescence microscopy shows that during NPC reassembly at the end of mitosis, Tpr becomes concentrated at the NE significantly later than O-linked glycoproteins, including p62. This indicates that reassembly of the NPC after mitosis is a stepwise process, and that the Tpr-containing peripheral structures are assembled later than p62.

Amino Acid Sequence↗
PubMed1993

[Suffocation attack].

An acute attack of suffocation is a life threatening event which may be fatal within minutes. Since the oxygen stores of the body are small, any interruption of oxygen uptake by the lungs results in immediate anoxia of tissue and in particular in perilous cerebral anoxia. The primary aims in the management of this emergency situation are: (1.) opening of the airways, clearing any obstruction, and keeping airways open; (2.) administration of oxygen and, if possible, assisted ventilation. This "reflex management" will gain time for diagnostic evaluation, for the decision about further management and, in most cases, for safe transport to the nearest emergency hospital. Further emergency measures are required in the case of upper airway obstruction due either to a foreign body or to laryngeal edema caused by local insect stings or anaphylaxis. Occasionally a foreign body obstructing the upper airways can be cleared using the Heimlich maneuver, i.e. rapid inward and upward pressure on the victim's epigastrium. Mild forms of laryngeal edema can be relieved by inhalation of epinephrine. If these methods fail, cricothyroidotomy or transtracheal ventilation must be performed. Patients with acute severe asthma must be immediately treated with inhalations of a beta-agonist, parenteral prednisolone and administration of oxygen, while emergency transport to the nearest hospital must be organized. To ensure first aid the emergency physician must be equipped with the necessary instruments such as a small oxygen cylinder, appropriate cannula with connectors and tubings for transtracheal ventilation, and a suitable bag-valve-mask device.

Airway Obstruction↗
PubMed1993

The rod domain of NF-L determines neurofilament architecture, whereas the end domains specify filament assembly and network formation.

Neurofilaments, assembled from NF-L, NF-M, and NF-H subunits, are the most abundant structural elements in myelinated axons. Although all three subunits contain a central, alpha-helical rod domain thought to mediate filament assembly, only NF-L self-assembles into 10-nm filaments in vitro. To explore the roles of the central rod, the NH2-terminal head and the COOH-terminal tail domain in filament assembly, full-length, headless, tailless, and rod only fragments of mouse NF-L were expressed in bacteria, purified, and their structure and assembly properties examined by conventional and scanning transmission electron microscopy (TEM and STEM). These experiments revealed that in vitro assembly of NF-L into bona fide 10-nm filaments requires both end domains: whereas the NH2-terminal head domain promotes lateral association of protofilaments into protofibrils and ultimately 10-nm filaments, the COOH-terminal tail domain controls lateral assembly of protofilaments so that it terminates at the 10-nm filament level. Hence, the two end domains of NF-L have antagonistic effects on the lateral association of protofilaments into higher-order structures, with the effect of the COOH-terminal tail domain being dominant over that of the NH2-terminal head domain. Consideration of the 21-nm axial beading commonly observed with 10-nm filaments, the approximate 21-nm axial periodicity measured on paracrystals, and recent cross-linking data combine to support a molecular model for intermediate filament architecture in which the 44-46-nm long dimer rods overlap by 1-3-nm head-to-tail, whereas laterally they align antiparallel both unstaggered and approximately half-staggered.

Animals↗
PubMed1992

[Intellectual and neurological development of 9-15-year-olds who were born as at-risk neonates].

This retrospective study deals with 296 children born between 1, Jan. 1974 and 31, Dec. 1980, who had been hospitalized as newborns at the Department of Pediatrics of the University of Berne and who had been considered at risk for abnormal psychomotor development because of well defined perinatal risk factors. Their psychomotor development had first been evaluated in the first months and years of life at the Cerebral Palsy Centre at Berne. In 1989, when the children were 9-15 years old, their neuro-intellectual development was investigated through a questionnaire sent to the families dealing with the subsequent psychosocial and scholastic course. In the CP-Centre, at a median age of 10 and mean age of 26.4 months, 247 (83.5%) of the children had been discharged with the finding of normal psychomotor development, 25 (8.4%) had been considered to show questionable findings and 24 (8.1%) to show obvious abnormal psychomotor development. On the basis of the answers to the questionnaire (median age 11 8/12, mean age 11 10/12 years; 62% return rate), 39.3% of the children had had no further problem, and 57.4% had used special support such as speech therapy or educational consulting and/or had had school problems. 3.3% showed distinct learning disabilities. More than 90% were attending a normal public school. This corresponds to the attendance in the general population. However, fewer children were in higher schools. To investigate whether the judgement in the first months and years (CP-Centre) of life corresponded to the further development (questionnaire), a subpopulation was specially evaluated in addition.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1992

2D crystallization: from art to science.

The techniques as well as the principles of the 2D crystallization of membrane and water-soluble proteins for electron crystallography are reviewed. First, the biophysics of the interactions between proteins, lipids and detergents is surveyed. Second, crystallization of membrane proteins in situ and by reconstitution methods is discussed, and the various factors involved are addressed. Third, we elaborate on the 2D crystallization of water-soluble proteins, both in solution and at interfaces, such as lipid monolayers, mica, carbon film or mercury surfaces. Finally, techniques and instrumentations that are required for 2D crystallization are described.

Aluminum Silicates↗
PubMed1992

Has negative staining still a place in biomacromolecular electron microscopy?

Transmission electron microscopy of proteins has provided molecular- and in a few cases near-atomic-resolution structural information. In this review, we critically evaluate the potential and the limitations in obtaining molecular resolution, particularly with negatively stained specimens, and put these into perspective with cryomicroscopy of unstained frozen-hydrated and sugar-embedded preparations.

Histocytological Preparation Techniques↗
PubMed1992

The structure of the F-actin filament and the actin molecule.

A consensus view on the three-dimensional structure of the F-actin filament and the relative strength of the intersubunit contacts in the filament has been established from an atomic filament model and recent three-dimensional reconstructions from electron micrographs of F-actin filaments. Functional implications of recent structural and biochemical data indicating a rather dynamic filament structure are discussed.

Actin Cytoskeleton↗