bluuebunny/crossref_metadata_2025_split huggingface dataset:
Sonication of proteins causes formation of aggregates that resemble amyloid
Authors: Stathopulos Peter B. , Scholz Guenter A. , Hwang Young‐Mi , Rumfeldt Jessica A.O. , Lepock James R. , Meiering Elizabeth M. 
Published: October 2004

AbstractDespite the widespread use of sonication in medicine, industry, and research, the effects of sonication on proteins remain poorly characterized. We report that sonication of a range of structurally diverse proteins results in the formation of aggregates that have similarities to amyloid aggregates. The formation of amyloid is associated with, and has been implicated in, causing of a wide range of protein conformational disorders including Alzheimer's disease, Huntington's disease, Parkinson's disease, and prion diseases. The aggregates cause large enhancements in fluorescence of the dye thioflavin T, exhibit green‐gold birefringence upon binding the dye Congo red, and cause a red‐shift in the absorbance spectrum of Congo red. In addition, circular dichroism reveals that sonication‐induced aggregates have high β‐content, and proteins with significant native α‐helical structure show increased β‐structure in the aggregates. Ultrastructural analysis by electron microscopy reveals a range of morphologies for the sonication‐induced aggregates, including fibrils with diameters of 5–20 nm. The addition of preformed aggregates to unsonicated protein solutions results in accelerated and enhanced formation of additional aggregates upon heating. The dye‐binding and structural characteristics, as well as the ability of the sonication‐induced aggregates to seed the formation of new aggregates are all similar to the properties of amyloid. These results have important implications for the use of sonication in food, biotechnological and medical applications, and for research on protein aggregation and conformational disorders.
Urea modulation of β‐amyloid fibril growth: Experimental studies and kinetic models
Authors: Kim Jin Ryoun , Muresan Adrian , Lee Ka Yee C. , Murphy Regina M. 
Published: October 2004

AbstractAggregation of β‐amyloid (Aβ) into fibrillar deposits is widely believed to initiate a cascade of adverse biological responses associated with Alzheimer's disease. Although it was once assumed that the mature fibril was the toxic form of Aβ, recent evidence supports the hypothesis that Aβ oligomers, intermediates in the fibrillogenic pathway, are the dominant toxic species. In this work we used urea to reduce the driving force for Aβ aggregation, in an effort to isolate stable intermediate species. The effect of urea on secondary structure, size distribution, aggregation kinetics, and aggregate morphology was examined. With increasing urea concentration, β‐sheet content and the fraction of aggregated peptide decreased, the average size of aggregates was reduced, and the morphology of aggregates changed from linear to a globular/linear mixture and then to globular. The data were analyzed using a previously published model of Aβ aggregation kinetics. The model and data were consistent with the hypothesis that the globular aggregates were intermediates in the amyloidogenesis pathway rather than alternatively aggregated species. Increasing the urea concentration from 0.4 M to 2 M decreased the rate of filament initiation the most; between 2 M and 4 M urea the largest change was in partitioning between the nonamyloid and amyloid pathways, and between 4 M and 6 M urea, the most significant change was a reduction in the rate of filament elongation.
Alteration of the tertiary structure of the major bee venom allergen Api m 1 by multiple mutations is concomitant with low IgE reactivity
Authors: Buhot Cécile , Chenal Alexandre , Sanson Alain , Pouvelle‐Moratille Sandra , Gelb Michael H. , Ménez André , Gillet Daniel , Maillère Bernard 
Published: October 2004

AbstractWe have engineered a recombinant form of the major bee venom allergen (Api m 1) with the final goal of reducing its IgE reactivity. This molecule (Api mut) contains 24 mutations and one deletion of 10 amino acids. The successive introduction of these sequence modifications led to a progressive loss of specific IgE and IgG reactivity and did not reveal any immunodominant epitopes. However, Api mut exhibited a clear loss of reactivity for Api m 1‐specific IgE and IgG. Injection of Api mut into mice induced specific antibody production. This humoral response was as high as that induced by the Api m 1 but the cross‐reactivity of the antibodies was weak. As inferred by far UV circular dichroism, this mutant was correctly folded. However, near UV circular dichroism and denaturation curves of Api mut showed that it exhibits a dynamic tertiary structure and that it is a highly flexible molecule. Finally, as all the sequence modifications have been introduced outside the human and murine T cell epitope regions, we investigated its T cell properties in mice. We showed that Api mut‐specific T lymphocytes induced in vivo were stimulated in vitro by both proteins. These data provide new insights in the design of hypoallergenic molecules.
Application of electrospray ionization mass spectrometry to study the hydrophobic interaction between the ε and θ subunits of DNA polymerase III
Authors: Gupta Rajesh , Hamdan Samir M. , Dixon Nicholas E. , Sheil Margaret M. , Beck Jennifer L. 
Published: October 2004

AbstractThe interactions between the N‐terminal domain of the ε (ε186) and θ subunits of DNA polymerase III of Escherichia coli were investigated using electrospray ionization mass spectrometry. The ε186–θ complex was stable in 9 M ammonium actetate (pH 8), suggesting that hydrophobic interactions have a predominant contribution to the stability of the complex. Addition of primary alkanols to ε186–θ in 0.1 M ammonium acetate (pH 8), led to dissociation of the complex, as observed in the mass spectrometer. The concentrations of methanol, ethanol, and 1‐propanol required to dissociate 50% of the complex were 8.9 M, 4.8 M, and 1.7 M, respectively. Closer scrutiny of the effect of alkanols on ε186, θ, and ε186–θ showed that ε186 formed soluble aggregates prior to precipitation, and that the association of ε186 with θ stabilized ε186. In‐source collision‐induced dissociation experiments and other results suggested that the ε186–θ complex dissociated in the mass spectrometer, and that the stability (with respect to dissociation) of the complex in vacuo was dependent on the solution from which it was sampled.
Insertion of the cytochrome b5 heme‐binding loop into an SH3 domain. Effects on structure and stability, and clues about the cytochrome's architecture
Authors: Knappenberger Jane A. , Kraemer‐Pecore Christina M. , Lecomte Juliette T.J. 
Published: October 2004

AbstractUnder native conditions, apocytochrome b5 exhibits a stable core and a disordered heme‐binding region that refolds upon association with the cofactor. The termini of this flexible region are in close proximity, suggesting that loop closure may contribute to the thermodynamic properties of the apocytochrome. A chimeric protein containing 43 residues encompassing the cytochrome loop was constructed using the cyanobacterial photosystem I accessory protein E (PsaE) from Synechococcus sp. PCC 7002 as a structured scaffold. PsaE has the topology of an SH3 domain, and the insertion was engineered to replace its 14‐residue CD loop. NMR and optical spectroscopies showed that the hybrid protein (named EbE1) was folded under native conditions and that it retained the characteristics of an SH3 domain. NMR spectroscopy revealed that structural and dynamic differences were confined near the site of loop insertion. Variable‐temperature 1D NMR spectra of EbE1 confirmed the presence of a kinetic unfolding barrier. Thermal and chemical denaturations of PsaE and EbE1 demonstrated cooperative, two‐state transitions; the stability of the PsaE scaffold was found only moderately compromised by the insertion, with a ΔTm of 8.3°C, a ΔCm of 1.5 M urea, and a ΔΔG° of 4.2 kJ/mole. The data implied that the penalty for constraining the ends of the inserted region was lower than the ∼6.4 kJ/mole calculated for a self‐avoiding chain. Extrapolation of these results to cytochrome b5 suggested that the intrinsic stability of the folded portion of the apoprotein reflected only a small detrimental contribution from the large heme‐binding domain.
Promiscuous protein biotinylation by Escherichia coli biotin protein ligase
Authors: Choi‐Rhee Eunjoo , Schulman Howard , Cronan John E. 
Published: October 2004

AbstractBiotin protein ligases (BPLs) are enzymes of extraordinary specificity. BirA, the BPL of Escherichia coli biotinylates only a single cellular protein. We report a mutant BirA that attaches biotin to a large number of cellular proteins in vivo and to bovine serum albumin, chloramphenicol acetyltransferase, immunoglobin heavy and light chains, and RNAse A in vitro. The mutant BirA also self biotinylates in vivo and in vitro. The wild type BirA protein is much less active in these reactions. The biotinylation reaction is proximity‐dependent in that a greater extent of biotinylation was seen when the mutant ligase was coupled to the acceptor proteins than when the acceptors were free in solution. This approach may permit facile detection and recovery of interacting proteins by existing avidin/streptavidin technology.
Mimicry by asx‐ and ST‐turns of the four main types of β‐turn in proteins
Authors: Duddy William J. , Nissink J. Willem M. , Allen Frank H. , Milner‐White E. James 
Published: October 2004

AbstractHydrogen‐bonded β‐turns in proteins occur in four categories: type I (the most common), type II, type II', and type I'. Asx‐turns resemble β‐turns, in that both have an NH…OC hydrogen bond forming a ring of 10 atoms. Serine and threonine side chains also commonly form hydrogen‐bonded turns, here called ST‐turns. Asx‐turns and ST‐turns can be categorized into four classes, based on side chain rotamers and the conformation of the central turn residue, which are geometrically equivalent to the four types of β‐turns. We propose asx‐ and ST‐turns be named using the type I, II, I', and II' β‐turn nomenclature. Using this, the frequency of occurrence of both asx‐ and ST‐turns is: type II' > type I > type II > type I', whereas for β‐turns it is type I > type II > type I' > type II'. Almost all type II asx‐turns occur as a recently described three residue feature named an asx‐nest.
Crystal structure of human coactosin‐like protein at 1.9 Å resolution
Authors: Li Xuemei , Liu Xueqi , Lou Zhiyong , Duan Xin , Wu Hao , Liu Yiwei , Rao Zihe 
Published: October 2004

AbstractHuman coactosin‐like protein (CLP) shares high homology with coactosin, a filamentous (F)‐actin binding protein, and interacts with 5LO and F‐actin. As a tumor antigen, CLP is overexpressed in tumor tissue cells or cell lines, and the encoded epitopes can be recognized by cellular and humoral immune systems. To gain a better understanding of its various functions and interactions with related proteins, the crystal structure of CLP expressed in Escherichia coli has been determined to 1.9 Å resolution. The structure features a central β‐sheet surrounded by helices, with two very tight hydrophobic cores on each side of the sheet. CLP belongs to the actin depolymerizing protein superfamily, and is similar to yeast cofilin and actophilin. Based on our structural analysis, we observed that CLP forms a polymer along the crystallographic b axis with the exact same repeat distance as F‐actin. A model for the CLP polymer and F‐actin binding has therefore been proposed.
Peroxidative aggregation of α‐synuclein requires tyrosines
Authors: Olteanu Alina , Pielak Gary J. 
Published: October 2004

Abstractα‐Synuclein is the main component of the intracellular protein aggregates in neurons of patients with Parkinson's disease. The occurrence of the disease is associated with oxidative damage. Although it is known that peroxidative chemistry leads to the aggregation of α‐synuclein in vitro, the specific amino acid types of α‐synuclein involved in this type of aggregation have not been identified. We show, using human cytochromecplus H2O2as the source oxidative stress, that the tyrosines of α‐synuclein are required for aggregation. The studies reveal the chemical basis for a crucial step in the aggregation process.
Crystal structure of a dodecameric FMN‐dependent UbiX‐like decarboxylase (Pad1) from Escherichia coli O157: H7
Authors: Rangarajan Erumbi S. , Li Yunge , Iannuzzi Pietro , Tocilj Ante , Hung Li‐Wei , Matte Allan , Cygler Miroslaw 
Published: October 2004

AbstractThe crystal structure of the flavoprotein Pad1 from Escherichia coli O157:H7 complexed with the cofactor FMN has been determined by the multiple anomalous diffraction method and refined at 2.0 Å resolution. This protein is a paralog of UbiX (3‐octaprenyl‐4‐hydroxybenzoate carboxylyase, 51% sequence identity) that catalyzes the third step in ubiquinone biosynthesis and to Saccharomyces cerevisiae Pad1 (54% identity), an enzyme that confers resistance to the antimicrobial compounds phenylacrylic acids through decarbox‐ylation of these compounds. Each Pad1 monomer consists of a typical Rossmann fold containing a non–covalently bound molecule of FMN. The fold of Pad1 is similar to MrsD, an enzyme associated with lantibiotic synthesis; EpiD, a peptidyl‐cysteine decarboxylase; and AtHAL3a, the enzyme, which decarboxylates 4′‐phosphopantothenoylcysteine to 4′‐phosphopantetheine during coenzyme A biosynthesis, all with a similar location of the FMN binding site at the interface between two monomers, yet each having little sequence similarity to one another. All of these proteins associate into oligomers, with a trimer forming the common structural unit in each case. In MrsD and EpiD, which belong to the homo‐dodecameric flavin‐containing cysteine decarboxylase (HFCD) family, these trimers associate further into dodecamers. Pad1 also forms dodecamers, although the association of the trimers is completely different, resulting in exposure of a different side of the trimer unit to the solvent. This exposure affects the location of the substrate binding site and, specifically, its access to the FMN cofactor. Therefore, Pad1 forms a separate family, distinguishable from the HFCD family.
Stabilization of membranes upon interaction of amphipathic polymers with membrane proteins
Authors: Picard Martin , Duval‐Terrié Caroline , Dé Emmanuelle , Champeil Philippe 
Published: October 2004

AbstractAmphipathic polymers derived from polysaccharides, namely hydrophobically modified pullulans, were previously suggested to be useful as polymeric substitutes of ordinary surfactants for efficient and structure‐conserving solubilization of membrane proteins, and one such polymer, 18C10, was optimized for solubilization of proteins derived from bacterial outer membranes (Duval‐Terrié et al. 2003). We asked whether a similar ability to solubilize proteins could also be demonstrated in eukaryotic membranes, namely sarcoplasmic reticulum (SR) fragments, the major protein of which is SERCA1a, an integral membrane protein with Ca2+‐dependent ATPase and Ca2+‐pumping activity. We found that 18C10‐mediated solubilization of these SR membranes did not occur. Simultaneously, however, we found that low amounts of this hydrophobically modified pullulan were very efficient at preventing long‐term aggregation of these SR membranes. This presumably occurred because the negatively charged polymer coated the membranous vesicles with a hydrophilic corona (a property shared by many other amphipathic polymers), and thus minimized their flocculation. Reminiscent of the old Arabic gum, which stabilizes Indian ink by coating charcoal particles, the newly designed amphipathic polymers might therefore unintentionally prove useful also for stabilization of membrane suspensions.
Is Good Policy Unimplementable? Reflections on the Ethnography of Aid Policy and Practice
Authors: Mosse David 
Published: October 2004

AbstractDespite the enormous energy devoted to generating the right policy models in development, strangely little attention is given to the relationship between these models and the practices and events that they are expected to generate or legitimize. Focusing on the unfolding activities of a development project over more than ten years as it falls under different policy regimes, this article challenges the assumption that development practice is driven by policy, suggesting that the things that make for ‘good policy’— policy which legitimizes and mobilizes political support — in reality make it rather unimplementable within its chosen institutions and regions. But although development practice is driven by a multi‐layered complex of relationships and the culture of organizations rather than policy, development actors work hardest of all to maintain coherent representations of their actions as instances of authorized policy, because it is always in their interest to do so. The article places these observations within the wider context of the anthropology of development and reflects on the place, method and contribution of development ethnography.
Resettlement, Rights to Development and the Ilisu Dam, Turkey
Authors: Morvaridi Behrooz 
Published: October 2004

AbstractA cursory attempt to measure the extent of displacement over the past two decades indicates significant increases in conflict‐induced displacement and displacement resulting from development projects. At the same time a growing opposition to the latter form of displacement has raised questions over its legitimacy through a variety of media, including public campaigns and protests. This article focuses on some of the challenges that this presents to the displacement and resettlement discourse. In particular it considers the influences of the rights to development agenda on the spatial context of displacement and its associated economic and political changes. There appears to be a disjuncture between the practices of mainstream development, which tend to interpret development policy as it is defined and applied by a nation state and to assess inequalities within clear geographical definitions, and the universality of a rights based approach to development. This article examines these tensions in the context of displacement and resettlement management, drawing on evidence from a case study of the Ilisu dam in South East Anatolia, Turkey.
Morphometric and immunohistochemical study of the reticulum of red deer during prenatal development
Authors: Franco A. J. , Redondo E. , Masot A. J. 
Published: October 2004

AbstractIberian red deer need to be conserved for their economic role and for their genetic importance as an important component of the ecosystem. Modifications currently being made to traditional management systems require a better understanding of the structure, function and development of their alimentary system. Here we describe a histomorphometric and immunohistochemical analysis of the stomach of 25 red deer embryos and fetuses from 30 days of gestation until birth (235 days). Differentiation of the reticular compartment from the primitive gastric tube begins at 67 days, forming a three‐layered structure: epithelium, pluripotential blastemal tissue and serosa. The primitive reticular cells are initiated as small epithelial evaginations (primary ribs) at 117 days. At 142 days, lateral growths appear from the primary reticular ribs, forming the corneum papillae. The secondary reticular ribs form at 142 days as growths from the primary ribs. The uneven height of primary and secondary reticular ribs leads to the formation of cells of varying size. Growth of the reticular ribs involves the lamina propria but not the submucosa, so clear separation of these layers is maintained during histodifferentiation. Formation of the tunica muscularis from the pluripotential blastemal tissue begins at 67 days of intrauterine life, as two layers of longitudinally and circularly arranged myoblasts. Differentiation of the muscularis from the mucosa occurs at approximately 205 days, as longitudinal projections of the internal bundles of the tunica muscularis form the musculature of the primary ribs. The secretion of neutral and acid mucopolysaccharides by the reticular epithelial layer begins at 67 days, establishing the gradual adaptation of the mucosa to its protective function in postnatal life. Neuroendocrine (non‐neuron enolase) and glial cells (glial fibrillary acidic protein and vimentin) were detected by immunohistochemistry, in a similar localization and intensity to that reported in the rumen. The neuropeptides vasoactive intestinal peptide and neuropeptide Y showed a positive immunoreaction in the reticular epithelium from 142 days of prenatal life, again earlier than reported for the rumen. In comparison with domestic ruminants, deer were shown to be less precocious with regard to development of gastric tube, in their capacity to secrete neutral mucopolysaccharides, and in their neuroendocrine nature, as determined by the detection of positive neuroedocrine and/or glial cells.
Tissue‐specific expression of the tight junction proteins claudins and occludin in the rat salivary glands
Authors: Peppi M. , Ghabriel M. N. 
Published: October 2004

AbstractTight junctions (TJs) are essential features of endothelial barrier membranes and of fluid‐secreting epithelial cells, such as in the salivary glands. Novel integral membrane proteins have been identified as components of TJs, namely claudins and occludin. The aim of the present study was to determine the distribution of occludin and claudins in the large salivary glands of the rat. The parotid, submandibular and sublingual salivary glands were harvested from adult Sprague–Dawley rats and cryostat sections were stained using immunoperoxidase and immunofluorescence methods. Claudin‐1 was expressed in endothelial cells of microvessels and in short selected segments of the duct system. Claudin‐3 was expressed principally in the acinar cells and intercalated ducts, while claudin‐4 was principally expressed by the striated and interlobular ducts. Claudin‐5 was specific to endothelial cells of microvessels. Occludin was ubiquitously detected in the duct system. Double labelling and confocal microscopy showed some co‐localization of claudin‐3 with claudin‐4, and minimal co‐localization of occludin with claudin‐4, in the striated ducts. Claudin 2 was not detected in any of the salivary glands. The results indicate specificity of the chemical composition of tight junctions in the rat salivary glands, and may reflect different physiological roles for TJs in the glandular and duct epithelial cells, and in endothelial cells of salivary gland microvessels.