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排序方式: 共有1005条查询结果,搜索用时 234 毫秒
71.
72.
González HE Eugenín EA Garcés G Solís N Pizarro M Accatino L Sáez JC 《American journal of physiology. Gastrointestinal and liver physiology》2002,282(6):G991-G1001
Hepatocyte gap junction proteins, connexins (Cxs) 26 and 32, are downregulated during obstructive cholestasis (OC) and lipopolysaccharide hepatocellular cholestasis (LPS-HC). We investigated rat hepatic Cxs during ethynylestradiol hepatocellular cholestasis (EE-HC) and choledochocaval fistula (CCF) and compared them with OC and LPS-HC. Levels (immunoblotting) and cellular distribution (immunofluorescence) of Cx26, -32, and -43, as well as macrophage infiltration, were studied in livers of rats under each condition. Cx26 and -32 were reduced in LPS-HC, OC, and CCF. However, in EE-HC, Cx26 did not change and Cx32 was increased. Prominent inflammation occurred in LPS-HC, OC, and CCF, which was associated with increased levels of Cx43 in LPS-HC and OC but not CCF. No inflammation nor changes in Cx43 levels occurred during EE-HC. In cultured hepatocytes, dye coupling was reduced by tumor necrosis factor-alpha and interleukins-1beta and -6, whereas reduction induced by LPS required coculture with Kupffer cells. Thus hepatocyte gap junctions are downregulated in forms of cholestasis associated with inflammation, and reduced intercellular communication might be induced in part by proinflammatory mediators. 相似文献
73.
Ohnishi T Yamashita A Kashima I Schell T Anders KR Grimson A Hachiya T Hentze MW Anderson P Ohno S 《Molecular cell》2003,12(5):1187-1200
Eukaryotic mRNAs containing premature termination codons (PTCs) are degraded by a process known as nonsense-mediated mRNA decay (NMD). NMD has been suggested to require the recognition of PTC by an mRNA surveillance complex containing UPF1/SMG-2. In multicellular organisms, UPF1/SMG-2 is a phosphoprotein, and its phosphorylation contributes to NMD. Here we show that phosphorylated hUPF1, the human ortholog of UPF1/SMG-2, forms a complex with human orthologs of the C. elegans NMD proteins SMG-5 and SMG-7. The complex also associates with protein phosphatase 2A (PP2A), resulting in dephosphorylation of hUPF1. Overexpression of hSMG-5 mutants that retain interaction with P-hUPF1 but which cannot induce its dephosphorylation impair NMD, suggesting that NMD requires P-hUPF1 dephosphorylation. We also show that P-hUPF1 forms distinct complexes containing different isoforms of hUPF3A. We propose that sequential phosphorylation and dephosphorylation of hUPF1 by hSMG-1 and PP2A, respectively, contribute to the remodeling of the mRNA surveillance complex. 相似文献
74.
We investigated the association between complement resistance and phenotypes of pathogenicity of Borrelia burgdorferi sensu lato isolates cultivated in a LEW/N rat tibiotarsal joint-derived tissue feeder layer-supported co-culture system. Guinea pig complement and immune serum raised in LHS/Ss hamsters caused complete lysis of B. burgdorferi sensu stricto isolate 297, B. afzelii and B. garinii in Barbour-Stoenner-Kelly's medium; however, tissue co-cultured B. burgdorferi sensu stricto contained complement escape variants. The arthritogenicity and infectivity of these variants were tested in 3-week-old Syrian hamsters and in a vaccinated hamster model in which formalin-killed B. burgdorferi sensu stricto C-1-11 vaccinated animals develop severe arthritis after challenge with live, pathogenic, low-passage 297 isolate. Non-animal-passaged complement escape variants were infectious in both animal models as demonstrated by re-isolation from the infected animals and competitive PCR. IP injection of animal-passaged complement escape variants caused development of severe arthritis in vaccinated animals 5 weeks post-injection; animal passage of complement escape variants was necessary for isolation of arthritogenic spirochetes from high-passaged, non-arthritogenic, attenuated borrelia cultures. Complement escape variants synthesized outer surface protein E as demonstrated by SDS-PAGE and western blotting analyses. The complement-mediated selection technique in tissue co-culture provides a novel approach to the studies of Lyme disease, enables us to isolate pathogenically distinct borrelia populations from attenuated cultures and prepare a moderately infectious, non-pathogenic live vaccine against this illness. 相似文献
75.
Blankenship JR Wormley FL Boyce MK Schell WA Filler SG Perfect JR Heitman J 《Eukaryotic cell》2003,2(3):422-430
76.
Tailoring host immune responses to Listeria by manipulation of virulence genes -- the interface between innate and acquired immunity 总被引:2,自引:0,他引:2
Peters C Domann E Darbouche A Chakraborty T Mielke ME 《FEMS immunology and medical microbiology》2003,35(3):243-253
Although attenuated strains of microbial pathogens have triggered vaccine development from its origin, the role of virulence factors in determining host immunity has remained largely unexplored. Using the murine listeriosis model, we investigated whether the induction and expansion of protective and inflammatory T cell responses may be modified by selective manipulation of virulence genes. We intentionally deleted specific genes of Listeria monocytogenes, including those encoding the positive regulatory factor (prfA), hemolysin (hly), the actin nucleator (actA), and phospholipase B (plcB). The resulting strains showed decisive differences in their immunogenic properties. In particular, we identified a double-deletion mutant that retained Listeria's profound ability to induce protective CD8(+) T cells, but that is strongly attenuated and exhibits a significantly reduced ability to induce CD4(+) T cell-mediated inflammation. We conclude that this mutant, L. monocytogenes DeltaactADeltaplcB, is at present the most promising mutant for a bacterial vaccine vector and is able to safely induce potent CD8(+) T cell-mediated immunity. 相似文献
77.
McCarthy DO Glowacki N Schell K Emler CA Albertini MR 《Cancer immunology, immunotherapy : CII》2000,49(2):85-93
The aim of this study was to compare the antigenicity of human melanoma cells molecularly modified by particle-mediated gene
transfer to have transient or stable expression of the B7-1 co-stimulatory molecule (CD80). The unmodified melanoma cells
(mel5, m21) had no constitutive expression of B7-1, but 22%–28% of cells had transient B7-1 expression 24 h following transfection
with cDNA for B7-1 (mel5-B7, m21-B7). In addition, 85%–90% of cells had stable B7-1 expression following transfection with
cDNA for B7-1 and in vitro culture under selection conditions (mel5-B7neo, m21-B7neo). Allogeneic HLA-unmatched normal donor
peripheral blood mononuclear cells (PBMC) secreted greater amounts of granulocyte/macrophage-colony-stimulating factor (GM-CSF)
when incubated for 3 days with m21-B7neo than did PBMC incubated with m21-B7, which, in turn, secreted greater amount of GM-CSF
than PBMC incubated with m21. Similarly, cell-mediated cytotoxicity against unmodified melanoma cells by PBMC co-cultured
for 5 days with the modified or unmodified melanoma cells was proportional to the level of B7-1 expression on the stimulating
cells. This cytolytic activity had both an HLA-class-I-restricted and an HLA-class-I-unrestricted component. Following 5 days
of co-culture, PBMC expression of CD28, the ligand for B7-1, was down-regulated in proportion to the level of B7-1 expression
on the stimulating melanoma cells. Thus, particle-mediated gene delivery of cDNA for B7-1 into human melanoma cells increased
expression of functional B7-1 and enhanced the antigenicity of the gene-modified cells in proportion to their level of B7-1
expression.
Received: 14 October 1999 / Accepted: 3 December 1999 相似文献
78.
Kajander T Kahn PC Passila SH Cohen DC Lehtiö L Adolfsen W Warwicker J Schell U Goldman A 《Structure (London, England : 1993)》2000,8(11):1203-1214
BACKGROUND: The traditional picture of charged amino acids in globular proteins is that they are almost exclusively on the outside exposed to the solvent. Buried charges, when they do occur, are assumed to play an essential role in catalysis and ligand binding, or in stabilizing structure as, for instance, helix caps. RESULTS: By analyzing the amount and distribution of buried charged surface and charges in proteins over a broad range of protein sizes, we show that buried charge is much more common than is generally believed. We also show that the amount of buried charge rises with protein size in a manner which differs from other types of surfaces, especially aromatic and polar uncharged surfaces. In large proteins such as hemocyanin, 35% of all charges are greater than 75% buried. Furthermore, at all sizes few charged groups are fully exposed. As an experimental test, we show that replacement of the buried D178 of muconate lactonizing enzyme by N stabilizes the enzyme by 4.2 degrees C without any change in crystallographic structure. In addition, free energy calculations of stability support the experimental results. CONCLUSIONS: Nature may use charge burial to reduce protein stability; not all buried charges are fully stabilized by a prearranged protein environment. Consistent with this view, thermophilic proteins often have less buried charge. Modifying the amount of buried charge at carefully chosen sites may thus provide a general route for changing the thermophilicity or psychrophilicity of proteins. 相似文献
79.
Substrate Specificity of and Product Formation by Muconate Cycloisomerases: an Analysis of Wild-Type Enzymes and Engineered Variants 总被引:3,自引:1,他引:2
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Martin Dominik Vollmer Helga Hoier Hans-Jürgen Hecht Ursula Schell Janosch Grning Adrian Goldman Michael Schlmann 《Applied microbiology》1998,64(9):3290-3299
Muconate cycloisomerases play a crucial role in the bacterial degradation of aromatic compounds by converting cis,cis-muconate, the product of catechol ring cleavage, to (4S)-muconolactone. Chloromuconate cycloisomerases catalyze both the corresponding reaction and a dehalogenation reaction in the transformation of chloroaromatic compounds. This study reports the first thorough examination of the substrate specificity of the muconate cycloisomerases from Pseudomonas putida PRS2000 and Acinetobacter “calcoaceticus” ADP1. We show that they transform, in addition to cis,cis-muconate, 3-fluoro-, 2-methyl-, and 3-methyl-cis,cis-muconate with high specificity constants but not 2-fluoro-, 2-chloro-, 3-chloro-, or 2,4-dichloro-cis,cis-muconate. Based on known three-dimensional structures, variants of P. putida muconate cycloisomerase were constructed by site-directed mutagenesis to contain amino acids found in equivalent positions in chloromuconate cycloisomerases. Some of the variants had significantly increased specificity constants for 3-chloro- or 2,4-dichloromuconate (e.g., A271S and I54V showed 27- and 22-fold increases, respectively, for the former substrate). These kinetic improvements were not accompanied by a change from protoanemonin to cis,cis-dienelactone as the product of 3-chloro-cis,cis-muconate conversion. The rate of 2-chloro-cis,cis-muconate turnover was not significantly improved, nor was this compound dehalogenated to any significant extent. However, the direction of 2-chloro-cis,cis-muconate cycloisomerization could be influenced by amino acid exchange. While the wild-type enzyme discriminated only slightly between the two possible cycloisomerization directions, some of the enzyme variants showed a strong preference for either (+)-2-chloro- or (+)-5-chloromuconolactone formation. These results show that the different catalytic characteristics of muconate and chloromuconate cycloisomerases are due to a number of features that can be changed independently of each other. 相似文献
80.
Characterization of Genes Involved in Biosynthesis of a Novel Antibiotic from Burkholderia cepacia BC11 and Their Role in Biological Control of Rhizoctonia solani 总被引:2,自引:0,他引:2
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Genetic manipulation of fluorescent pseudomonads has provided major insight into their production of antifungal molecules and their role in biological control of plant disease. Burkholderia cepacia also produces antifungal activities, but its biological control activity is much less well characterized, in part due to difficulties in applying genetic tools. Here we report genetic and biochemical characterization of a soil isolate of B. cepacia relating to its production of an unusual antibiotic that is very active against a variety of soil fungi. Purification and preliminary structural analyses suggest that this antibiotic (called AFC-BC11) is a novel lipopeptide associated largely with the cell membrane. Analysis of conditions for optimal production of AFC-BC11 indicated stringent environmental regulation of its synthesis. Furthermore, we show that production of AFC-BC11 is largely responsible for the ability of B. cepacia BC11 to effectively control the damping-off of cotton caused by the fungal pathogen Rhizoctonia solani in a gnotobiotic system. Using Tn5 mutagenesis, we identified, cloned, and characterized a region of the genome of strain BC11 that is required for production of this antifungal metabolite. DNA sequence analysis suggested that this region encodes proteins directly involved in the production of a nonribosomally synthesized lipopeptide. 相似文献