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51.
Henry C. Orr Kyle H. Sibinovic Peter G. Probst H. Donald Hochstein Dewitt C. Littlejohn 《In vitro cellular & developmental biology. Plant》1975,11(4):230-233
Summary Bacteriological tests were made on 24 lots of unfiltered calf serum collected for subsequent use as a component of tissue
culture media. The examination included the isolation and identification of bacteria, assay of phages, and demonstration of
endotoxin material. Only Gram-positive bacteria were isolated and 96% of the sera were contaminated with bacteria. The prevalent
strains of bacteria found wereBacillus species and streptococci and 63% of the sera coagulatedLimulus amebocyte lysate. More than 90% of the lots contained phages demonstrable with the C-3000 strain ofEscherichia coli. Only one lot of the serum was found to be free from bacteria, phages, and endotoxin by the tests used. 相似文献
52.
A technique is reported for cryostat sectioning of lung tissue in an expanded state for use in viral immunofluorescence studies. A 1: 2 mixture of O.C.T. embedding compound and phosphate-buffered saline is injected intratracheally into fresh lung tissne. The lung tissue is frozen in liquid nitrogen and sectioned with a cryostat. Compared to other published reports of lung sectioning for immunofluorescence miscroscopy, this method has the advantages of bekg easy and quick, maintaining the lung sectiom in an expanded rather than collapsed state and avoiding contact with chemicals potentially capable of altering sensitive viral antigens. 相似文献
53.
Joshua C Bufton Kyle T Powers Jenn-Yeu A Szeto Christine Toelzer Imre Berger Christiane Schaffitzel 《Nucleic acids research》2022,50(10):5934
UPF3 is a key nonsense-mediated mRNA decay (NMD) factor required for mRNA surveillance and eukaryotic gene expression regulation. UPF3 exists as two paralogs (A and B) which are differentially expressed depending on cell type and developmental stage and believed to regulate NMD activity based on cellular requirements. UPF3B mutations cause intellectual disability. The underlying molecular mechanisms remain elusive, as many of the mutations lie in the poorly characterized middle-domain of UPF3B. Here, we show that UPF3A and UPF3B share structural and functional homology to paraspeckle proteins comprising an RNA-recognition motif-like domain (RRM-L), a NONA/paraspeckle-like domain (NOPS-L), and extended α-helical domain. These domains are essential for RNA/ribosome-binding, RNA-induced oligomerization and UPF2 interaction. Structures of UPF2′s third middle-domain of eukaryotic initiation factor 4G (MIF4GIII) in complex with either UPF3B or UPF3A reveal unexpectedly intimate binding interfaces. UPF3B’s disease-causing mutation Y160D in the NOPS-L domain displaces Y160 from a hydrophobic cleft in UPF2 reducing the binding affinity ∼40-fold compared to wildtype. UPF3A, which is upregulated in patients with the UPF3B-Y160D mutation, binds UPF2 with ∼10-fold higher affinity than UPF3B reliant mainly on NOPS-L residues. Our characterization of RNA- and UPF2-binding by UPF3′s middle-domain elucidates its essential role in NMD. 相似文献
54.
55.
Purification and characterization of a phosphatidylinositol 4-kinase from bovine uteri 总被引:4,自引:0,他引:4
Phosphatidylinositol 4-kinase has been purified 10,148-fold to a specific activity of 2.7 mumol/mg/min from bovine uteri. This purification was accomplished by detergent extraction of an acetone powder, ammonium sulfate precipitation, and chromatography on MonoQ, S-Sepharose, MonoP, and hydroxylapatite columns. The purified enzyme has a molecular mass of 55 kDa and appears to be monomeric. Kinetic analyses of the enzymatic activity demonstrated apparent Km values of 18 microM and 22 micrograms/ml (approximately 26 microM) for ATP and phosphatidylinositol, respectively, optimal activity in the pH range of 6.0-7.0, and a sigmoidal dependence of enzymatic activity on [Mg2+]. Ca2+ inhibited the enzyme at nonphysiological concentrations with 50% inhibition observed at a free [Ca2+] of approximately 300 microM. The purified enzyme efficiently utilized both ATP and 2'-deoxy-ATP as phosphoryl donors and specifically phosphorylated phosphatidylinositol on the fourth position. No phosphatidylinositol-4-phosphate 5-kinase activity was observed in the purified enzyme preparations. To our knowledge, this is the first reported purification of a phosphatidylinositol-specific phosphatidylinositol 4-kinase. 相似文献
56.
57.
The renal toxicity of (R,S)-3-chlorolactate has been shown to be due to the (R)-isomer which, when administered to rats, induces diuresis and glucosuria. The metabolic activity of isolated tubule cells, prepared from rat kidney, was inhibited by (R)-3-chlorolactate and the action of the compound was localised as affecting mitochondrial metabolism. Studies with kidney mitochondria pin-pointed the site of action as being involved with the oxidative metabolism of malate but not the inhibition of mitochondrial malate dehydrogenase. The effects of oxalate, a metabolite of (R)-3-chlorolactate, and of (R,S)-3-chlorolactaldehyde on renal tubule cells was investigated. While some degrees of inhibition of metabolic activity were evident, these compounds were not responsible for the toxic effects produced by (R)-3-chlorolactate. 相似文献
58.
Lombardi ML Jaalouk DE Shanahan CM Burke B Roux KJ Lammerding J 《The Journal of biological chemistry》2011,286(30):26743-26753
Maintaining physical connections between the nucleus and the cytoskeleton is important for many cellular processes that require coordinated movement and positioning of the nucleus. Nucleo-cytoskeletal coupling is also necessary to transmit extracellular mechanical stimuli across the cytoskeleton to the nucleus, where they may initiate mechanotransduction events. The LINC (Linker of Nucleoskeleton and Cytoskeleton) complex, formed by the interaction of nesprins and SUN proteins at the nuclear envelope, can bind to nuclear and cytoskeletal elements; however, its functional importance in transmitting intracellular forces has never been directly tested. This question is particularly relevant since recent findings have linked nesprin mutations to muscular dystrophy and dilated cardiomyopathy. Using biophysical assays to assess intracellular force transmission and associated cellular functions, we identified the LINC complex as a critical component for nucleo-cytoskeletal force transmission. Disruption of the LINC complex caused impaired propagation of intracellular forces and disturbed organization of the perinuclear actin and intermediate filament networks. Although mechanically induced activation of mechanosensitive genes was normal (suggesting that nuclear deformation is not required for mechanotransduction signaling) cells exhibited other severe functional defects after LINC complex disruption; nuclear positioning and cell polarization were impaired in migrating cells and in cells plated on micropatterned substrates, and cell migration speed and persistence time were significantly reduced. Taken together, our findings suggest that the LINC complex is critical for nucleo-cytoskeletal force transmission and that LINC complex disruption can result in defects in cellular structure and function that may contribute to the development of muscular dystrophies and cardiomyopathies. 相似文献
59.
Wild ruminants are susceptible to infection from generalist helminth species, which can also infect domestic ruminants. A better understanding is required of the conditions under which wild ruminants can act as a source of helminths (including anthelmintic-resistant genotypes) for domestic ruminants, and vice versa, with the added possibility that wildlife could act as refugia for drug-susceptible genotypes and hence buffer the spread and development of resistance. Helminth infections cause significant productivity losses in domestic ruminants and a growing resistance to all classes of anthelmintic drug escalates concerns around helminth infection in the livestock industry. Previous research demonstrates that drug-resistant strains of the pathogenic nematode Haemonchus contortus can be transmitted between wild and domestic ruminants, and that gastro-intestinal nematode infections are more intense in wild ruminants within areas of high livestock density. In this article, the factors likely to influence the role of wild ruminants in helminth infections and anthelmintic resistance in livestock are considered, including host population movement across heterogeneous landscapes, and the effects of climate and environment on parasite dynamics. Methods of predicting and validating suspected drivers of helminth transmission in this context are considered based on advances in predictive modelling and molecular tools. 相似文献
60.
Nantermet PG Burgey CS Robinson KA Pellicore JM Newton CL Deng JZ Selnick HG Lewis SD Lucas BJ Krueger JA Miller-Stein C White RB Wong B McMasters DR Wallace AA Lynch JJ Yan Y Chen Z Kuo L Gardell SJ Shafer JA Vacca JP Lyle TA 《Bioorganic & medicinal chemistry letters》2005,15(11):2771-2775
In this study, we have demonstrated that the critical hydrogen bonding motif of the established 3-aminopyrazinone thrombin inhibitors can be effectively mimicked by a 2-aminopyridine N-oxide. As this peptidomimetic core is more resistant toward oxidative metabolism, it also overcomes the metabolic liability associated with the pyrazinones. An optimization study of the P(1) benzylamide delivered the potent thrombin inhibitor 21 (K(i) = 3.2 nM, 2xaPTT = 360 nM), which exhibited good plasma levels and half-life after oral dosing in the dog (C(max) = 2.6 microM, t(1/2) = 4.5 h). 相似文献