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991.
Transmissible spongiform encephalopathies are a class of fatal neurodegenerative diseases linked to the prion protein. The prion protein normally exists in a soluble, globular state (PrP(C)) that appears to participate in copper metabolism in the central nervous system and/or signal transduction. Infection or disease occurs when an alternatively folded form of the prion protein (PrP(Sc)) converts soluble and predominantly alpha-helical PrP(C) into aggregates rich in beta-structure. The structurally disordered N-terminus adopts beta-structure upon conversion to PrP(Sc) at low pH. Chemical chaperones, such as trimethylamine N-oxide (TMAO), can prevent formation of PrP(Sc) in scrapie-infected mouse neuroblastoma cells [Tatzelt, J., et al. (1996) EMBO J. 15, 6363-6373]. To explore the mechanism of TMAO protection of PrP(C) at the atomic level, molecular dynamics simulations were performed under conditions normally leading to conversion (low pH) with and without 1 M TMAO. In PrP(C) simulations at low pH, the helix content drops and the N-terminus is brought into the small native beta-sheet, yielding a PrP(Sc)-like state. Addition of 1 M TMAO leads to a decreased radius of gyration, a greater number of protein-protein hydrogen bonds, and a greater number of tertiary contacts due to the N-terminus forming an Omega-loop and packing against the structured core of the protein, not due to an increase in the level of extended structure as with the PrP(C) to PrP(Sc) simulation. In simulations beginning with the "PrP(Sc)-like" structure (derived from PrP(C) simulated at low pH in pure water) in 1 M TMAO, similar structural reorganization at the N-terminus occurred, disrupting the extended sheet. The mechanism of protection by TMAO appears to be exclusionary in nature, consistent with previous theoretical and experimental studies. The TMAO-induced N-terminal conformational change prevents residues that are important in the conversion of PrP(C) to PrP(Sc) from assuming extended sheet structure at low pH. 相似文献
992.
Baquero MR Nilsson AI Turrientes Mdel C Sandvang D Galán JC Martínez JL Frimodt-Møller N Baquero F Andersson DI 《Journal of bacteriology》2004,186(16):5538-5542
Polymorphisms in the rifampin resistance mutation frequency (f) were studied in 696 Escherichia coli strains from Spain, Sweden, and Denmark. Of the 696 strains, 23% were weakly hypermutable (4 x 10(-8) < or = f < 4 x 10(-7)), and 0.7% were strongly hypermutable (f > or = 4 x 10(-7)). Weak mutators were apparently more frequent in southern Europe and in blood isolates (38%) than in urinary tract isolates (25%) and feces of healthy volunteers (11%). 相似文献
993.
Ruiz de Almodóvar C Ruiz-Ruiz C Rodríguez A Ortiz-Ferrón G Redondo JM López-Rivas A 《The Journal of biological chemistry》2004,279(6):4093-4101
Tumor necrosis factor-related apoptosis-inducing ligand receptor 3 (TRAIL-R3) is a decoy receptor for TRAIL, a member of the tumor necrosis factor family. In several cell types decoy receptors inhibit TRAIL-induced apoptosis by binding TRAIL and thus preventing its binding to proapoptotic TRAIL receptors. We studied the regulation of TRAIL-R3 gene expression in breast tumor cells treated with the genotoxic drug doxorubicin (DXR). The breast tumor cell line MCF-7 (p53 wild type) responded to DXR with a marked elevation of TRAIL-R3 expression at the mRNA, total protein, and cell surface levels. In contrast, in EVSA-T cells (p53 mutant) DXR did not induce increased expression of TRAIL-R3. In MCF-7 cells overexpressing the human papillomavirus protein E6, which causes p53 degradation, DXR-induced TRAIL-R3 expression was notably reduced. Furthermore, in MCF-7 cells overexpressing a temperature-sensitive p53 mutant (Val135), shifting the cultures to the permissive temperature was sufficient to induce the expression of TRAIL-R3. We also cloned and characterized a p53 consensus element located within the first intron of the human TRAIL-R3 gene. This element binds p53 and confers responsiveness to genotoxic damage to constructs of the TRAIL-R3 promoter in transient transfection experiments. Our results indicate that genotoxic treatments such as DXR, frequently used in cancer therapy, may also induce genes such as TRAIL-R3 that potentially have antiapoptotic actions and thus interfere with the TRAIL signaling system. This is particularly important in view of the proposed use of TRAIL in antitumor therapy. 相似文献
994.
Rab2 requires atypical protein kinase C iota/lambda (aPKC iota/lambda) to promote vesicle formation from vesicular tubular clusters (VTCs). The Rab2-generated vesicles are enriched in recycling proteins suggesting that the carriers are retrograde-directed and retrieve transport machinery back to the endoplasmic reticulum. These vesicles also contained the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH). We have previously established that GAPDH is required for membrane transport between the endoplasmic reticulum and the Golgi complex. Moreover, GAPDH is phosphorylated by aPKC iota/lambda and binds to the aPKC iota/lambda regulatory domain. In this study, we employed a combination of in vivo and in vitro assays and determined that GAPDH also interacts with Rab2. The site of GAPDH interaction was mapped to Rab2 residues 20-50. In addition to its glycolytic function, GAPDH has multiple intracellular roles. However, the function of GAPDH in the early secretory pathway is unknown. One possibility is that GAPDH ultimately provides energy in the form of ATP. To determine whether GAPDH catalytic activity was critical for transport in the early secretory pathway, a conservative substitution was made at Cys-149 located at the active site, and the mutant was biochemically characterized in a battery of assays. Although GAPDH (C149G) has no catalytic activity, Rab2 recruited the mutant protein to membranes in a quantitative binding assay. GAPDH (C149G) is phosphorylated by aPKC iota/lambda and binds directly to Rab2 when evaluated in an overlay binding assay. Importantly, VSV-G transport between the ER and Golgi complex is restored when an in vitro trafficking assay is performed with GAPDH-depleted cytosol and GAPDH (C149G). These data suggest that GAPDH imparts a unique function necessary for membrane trafficking from VTCs that does not require GAPDH glycolytic activity. 相似文献
995.
The thallus formed by the marine pyrenomycete fungus Verrucaria tavaresiae and the phaeophycean alga Petroderma maculiforme was studied to elucidate the organization of the symbionts, determine the type of cellular contacts between them, and evaluate the status of the symbiosis as a lichen. Hand-sectioned and resin-embedded samples were examined with light and transmission electron microscopy. Within the uppermost portion of the cellular fungal tissue, separate algal filaments were arranged anticlinally. Protrusions of the fungal cell wall penetrated into adjacent algal walls but did not enter the cell lumen. A striking feature of these penetrations was the frequent separation of algal cell wall layers and insertion of fungal wall material between them. Algal filaments grew downward intrusively between fungal cells, often penetrating deeply into the fungal cell wall. Despite the exceptional nature of the phycobiont involved, the Verrucaria tavaresiae-Petroderma maculiforme symbiosis unequivocally fits the prevailing concept of a lichen. The distinctive interpenetrations observed between symbionts may be related to the integration of their different growth forms within a coherent tissue regularly subject to mechanical stresses. Periclinal cell divisions within and just below the algal layer may serve to replenish surface tissues lost to abrasion and herbivory. 相似文献
996.
Nonmuscle myosin heavy-chain gene MYH14 is expressed in cochlea and mutated in patients affected by autosomal dominant hearing impairment (DFNA4) 总被引:7,自引:0,他引:7 下载免费PDF全文
Donaudy F Snoeckx R Pfister M Zenner HP Blin N Di Stazio M Ferrara A Lanzara C Ficarella R Declau F Pusch CM Nürnberg P Melchionda S Zelante L Ballana E Estivill X Van Camp G Gasparini P Savoia A 《American journal of human genetics》2004,74(4):770-776
Myosins have been implicated in various motile processes, including organelle translocation, ion-channel gating, and cytoskeleton reorganization. Different members of the myosin superfamily are responsible for syndromic and nonsyndromic hearing impairment in both humans and mice. MYH14 encodes one of the heavy chains of the class II nonmuscle myosins, and it is localized within the autosomal dominant hearing impairment (DFNA4) critical region. After demonstrating that MYH14 is highly expressed in mouse cochlea, we performed a mutational screening in a large series of 300 hearing-impaired patients from Italy, Spain, and Belgium and in a German kindred linked to DFNA4. This study allowed us to identify a nonsense and two missense mutations in large pedigrees, linked to DFNA4, as well as a de novo allele in a sporadic case. Absence of these mutations in healthy individuals was tested in 200 control individuals. These findings clearly demonstrate the role of MYH14 in causing autosomal dominant hearing loss and further confirm the crucial role of the myosin superfamily in auditive functions. 相似文献
997.
L-type Ca2+ channels in Ca2+ channelopathies 总被引:3,自引:0,他引:3
Striessnig J Hoda JC Koschak A Zaghetto F Müllner C Sinnegger-Brauns MJ Wild C Watschinger K Trockenbacher A Pelster G 《Biochemical and biophysical research communications》2004,322(4):1341-1346
Voltage-gated L-type Ca2+ channels (LTCCs) mediate depolarization-induced Ca2+ entry in electrically excitable cells, including muscle cells, neurons, and endocrine and sensory cells. In this review we summarize the role of LTCCs for human diseases caused by genetic Ca2+ channel defects (channelopathies). LTCC dysfunction can result from structural aberrations within pore-forming alpha1 subunits causing incomplete congenital stationary night blindness, malignant hyperthermia sensitivity or hypokalemic periodic paralysis. However, studies in mice revealed that LTCC dysfunction also contributes to neurological symptoms in Ca2+ channelopathies affecting non-LTCCs, such as Ca(v)2.1 alpha1 in tottering mice. Ca2+ channelopathies provide exciting molecular tools to elucidate the contribution of different LTCC isoforms to human diseases. 相似文献
998.
Ylianttila MS Qin YM Hiltunen JK Glumoff T 《Biochemical and biophysical research communications》2004,324(1):25-30
The N-terminal part of Candida tropicalis MFE-2 (MFE-2(h2Delta)) having two (3R)-hydroxyacyl-CoA dehydrogenases with different substrate specificities has been purified and crystallized as a recombinant protein. The expressed construct was modified so that a stabile, homogeneous protein could be obtained instead of an unstabile wild-type form with a large amount of cleavage products. Cubic crystals with unit cell parameters a=74.895, b=78.340, c=95.445, and alpha=beta=gamma=90 degrees were obtained by using PEG 4000 as a precipitant. The crystals exhibit the space group P2(1)2(1)2(1) and contain one molecule, consisting of two different (3R)-hydroxyacyl-CoA dehydrogenases, in the asymmetric unit. The crystals diffract to a resolution of 2.2A at a conventional X-ray source. 相似文献
999.
Chambers SJ Wickham MS Regoli M Bertelli E Gunning PA Nicoletti C 《Biochemical and biophysical research communications》2004,325(4):1258-1263
Although the route of sensitization to food allergens is still the subject of debate, it is generally accepted the gut immune system plays a pivotal role. However, hitherto the transport of allergens across the normal, pre-sensitized gut epithelium remained largely unknown. Our aim was to identify the route through which protein bodies and soluble proteins from digested peanuts penetrated the pre-sensitized gut epithelium in vivo and the specific cell types involved in the transport. Digestion of peanuts released a large number of protein bodies that are exclusively transported across the epithelium by specialized antigen-sampling M cells and delivered to the lymphoid tissue of Peyer's patch. Intracellular transport of soluble protein also occurred almost exclusively via M cells and it was negligible across absorptive enterocytes. We hypothesize that these conditions which are known to favour strongly the induction of immune responses rather than oral tolerance may play a significant role in the genesis of allergic reactions. 相似文献
1000.
Díaz BR Abrahantes-Pérez Mdel C González M Valdés R Pérez ME 《Biochemical and biophysical research communications》2004,317(2):614-617
In this paper we describe an immunoenzymatic assay based on a rat monoclonal antibody (Ram kappa) developed to determine mouse IgG concentration, which is widely used for samples obtained on purification processes, like supernatant waste and the content of IgG in the vaccine (rHBsAg). This assay involves the use of a rat antibody-horseradish peroxidase-conjugated for the revealing of the antigen-antibody reaction. The rat antibody was produced in cell culture using a dialysis tube (DT). The immunoassay was standardized following several concepts, such as specificity, precision, and linearity. The result obtained permitted us to replace the use of polyclonal antibodies to determine the kappa light chain mouse antibodies by a rat monoclonal antibody of high sensibility and reproducibility. The assay permitted a reliable measurement of murine kappa Ig up to 0.68 ng/ml and was capable of quantifying 6.25 ng/ml. Due to the high frequency of the kappa light chain in mouse antibodies this system acquires a great application. 相似文献