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D S Schmid 《Journal of immunology (Baltimore, Md. : 1950)》1988,140(10):3610-3616
The nature of the in vitro human cytotoxic T-cell responder population to HSV type 1 (HSV-1) was studied. In 5-day HSV-1-stimulated cultures that contained MHC-restricted activity, two phenotypically distinct populations of cells were present that were capable of lysing HSV-1-infected B cell lines in a 5-h 51Cr-release assay. The first was CD4+, CD8-, CD16- cell typical of class II-restricted T cells, whereas the other population bore a CD4-, CD8-, CD16+ NK-cell phenotype. Elimination of the NK cell fraction from bulk cultures by using anti-CD16 plus C frequently resulted in cell populations that killed in an Ag-specific, HLA-DR-restricted fashion. In some cases the anti-CD16-pretreated cultures retained a killing population that was unrestricted to MHC products. In no instance were any cytotoxic T cells that were restricted to class I Ag in evidence. Limiting dilution analysis of precursor frequency indicated that about 1 in 4000 to 1 in 8000 cells from peripheral blood are specific for HSV-1 in seropositive individuals. Comparisons of HLA class I-matched and HLA class II-matched targets with the autologous target by using limiting dilution analysis yielded results entirely consistent with those obtained in the bulk culture assay system. 相似文献
14.
Rat sciatic nerve cytosol contains a phosphodiesterase of the phospholipase C type that catalyzes the hydrolysis of inositol phospholipids, with preferences of phosphatidylinositol 4'-phosphate (PIP) greater than phosphatidylinositol (PI) much greater than phosphatidylinositol 4',5'-bisphosphate (PIP2), at a pH optimum of 5.5-6.0 and at maximum rates of 55, 13, and 0.7 nmol/min/mg protein, respectively. Analysis of reaction products by TLC and formate exchange chromatography shows that inositol 1,2-cyclic phosphate (83%) and diacylglycerol are the major products of PI hydrolysis. [32P]-PIP hydrolysis yields inositol bisphosphate, inositol phosphate, and inorganic phosphate, indicating the presence of phosphodiesterase, phosphomonoesterase, and/or inositol phosphate phosphatase activities in nerve cytosol. Phosphodiesterase activity is Ca2+-dependent and completely inhibited by EGTA, but phosphomonoesterase activity is independent of divalent cations or chelating agents. Phosphatidylcholine (PC) and lysophosphatidylcholine (lysoPC) inhibit PI hydrolysis. They stimulate PIP and PIP2 hydrolysis up to equimolar concentrations, but are inhibitory at higher concentrations. Both diacylglycerols and free fatty acids stimulate PI hydrolysis and counteract its inhibition by PC and lysoPC. PIP2 is a poor substrate for the cytosolic phospholipase C and strongly inhibits hydrolysis of PI. However, it enhances PIP hydrolysis up to an equimolar concentration. 相似文献
15.
F0 portion of Escherichia coli ATP synthase: orientation of subunit c in the membrane 总被引:3,自引:0,他引:3
Incubation of right-side-out oriented membrane vesicles of Escherichia coli with tetranitromethane resulted in the nitration of tyrosine residues (Tyr-10 and Tyr-73) of subunit c from the ATP synthase. Cleavage of the protein with cyanogen bromide and separation of the resulting fragments, especially of the tyrosine-containing peptides, clearly demonstrated that the distribution of the nitro groups is similar at any time and at any pH value chosen for the analysis. Furthermore, the percentage of 3-nitrotyrosine present in the two peptide fragments was in good agreement with that obtained for the intact polypeptide chain. While the modification of the tyrosine residues in subunit c with the lipophilic tetranitromethane is independent of the orientation of the membrane vesicles, the subsequent partial conversion of the 3-nitrotyrosine to the amino form only occurred when membrane vesicles with right-side-out orientation were treated with the ionic, water-soluble sodium dithionite, which at certain concentrations cannot penetrate biological membranes. Cleavage of subunit c isolated from nitrated and subsequently reduced membrane vesicles and separation of the resulting fragments by high-pressure liquid chromatography showed that the 3-nitrotyrosine in the Tyr-73-containing peptides has been completely reduced, while the nitro group in peptides containing Tyr-10 remained nearly unaffected. 相似文献
16.
Dog heart microsomes catalyze the transfer of acyl groups from the sn-2 position of phosphatidylcholine (PC) to lysophosphatidylserine (lysoPS) in the presence of coenzyme A (CoA) at pH optima of 4.5-5.0 and 7.5. Acyl transfer activity at acidic pH is about three times higher than at neutral pH. Transacylation of lysoPS by acyl transfer from PC with dog heart microsomes at neutral pH favors arachidonate over linoleate by a factor of 2.1, whereas free linoleic acid is favored by a factor of 3.7 over arachidonic acid for lysoPS acylation in the presence of acyl-CoA-generating cofactors. Considering the location and acyl composition of myocardial PS, it appears that both acyl transfer from PC and utilization of unesterified fatty acids may be involved in the acylation of lysoPS at its sn-2 position. 相似文献
17.
A beta-glucosidase (E.C. 3.2.1.21) was isolated from the culture filtrate of fungus Trichoderma reesei QM 9414 grown in continuous culture with biomass retention. The crude extracellular enzyme preparation was fractionated by a three-step purification procedure [chromatography on Fractogel HW-55 (S) and Bio-Gel A 0.5 plus final preparative isoelectric focusing] to yield three beta-glucosidases with isoelectric points at pH 8.4, 8.0, and 7.4. Only one enzyme (pi 8.4) met the stringent criterion of being homogeneous according to titration curve analysis. This enzyme was then characterized not to be a glycoprotein, although the native protein contained 35% carbohydrate (as glucose). It was found to have an apparent molar mass of 7 x 10(4) g/mol (SDS-PAGE), exhibited its optimum activity towards cellobiose at pH 4.5 and 70 degrees C (30 min test), and lost less than 3% activity at 50 degrees C over a period of 7 h. The K(M) values towards cellobiose and p-nitrophenyl-beta-D-glucopyranoside were determined to be 0.5mM and 0.3mM, respectively. The enzyme hydrolyzed cellodextrins (cellotriose to cellooctaose) by sequentially splitting off glucose units from the nonreducing end of the oligomers. The extent of the observed transfer reactions varied with the initial substrate concentration. No enzyme activity towards microcrystalline cellulose or carboxymethylcellulose could be detected. The classification of the enzyme as beta-glucosidase or exo-beta-1,4-glucan glucohydrolase is discussed with respect to the exhibited hydrolytic activities. 相似文献
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19.
S. Kauffer R. Schmid K. Steffens G. Deckers-Hebestreit K. Altendorf 《Archives of microbiology》1987,148(3):187-192
The ATP synthase complex of Klebsiella pneumoniae (KF1F0) has been purified and characterized. SDS-gel electrophoresis of the purified F1F0 complexes revealed an identical subunit pattern for E. coli (EF1F0) and K. pneumoniae. Antibodies raised against EF1 complex and purified EF0 subunits recognized the corresponding polypeptides of EF1F0 and KF1F0 in immunoblot analysis. Protease digestion of the individual subunits generated an identical cleavage pattern for subunits , , , , a, and c of both enzymes. Only for subunit different cleavage products were obtained. The isolated subunit c of both organisms showed only a slight deviation in the amino acid composition. These data suggest that extensive homologies exist in primary and secondary structure of both ATP synthase complexes reflecting a close phylogenetic relationship between the two enterobacteric tribes.Abbreviations ACMA
9-amino-6-chloro-2-methoxyacridine
- DCCD
N,N-dicyclohexylcarbodiimide
- FITC
fluorescein isothiocyanate
- SDS
sodium dodecyl sulfate
- TTFB
4,5,6,7-tetrachloro-2-trifluoromethylbenzimidazole 相似文献
20.
Cytogenetic effects of 3,4-dichloroaniline in human lymphocytes and V79 Chinese hamster cells 总被引:2,自引:0,他引:2
3,4-Dichloroaniline (3,4-DCA), an intermediate in various chemical syntheses, has been detected as an environmental contaminant in surface waters and in the effluents from dye-manufacturing plants. Tested for clastogenicity in human lymphocytes in vitro the compound was inactive in the chromosome aberration assay yet exhibited a positive sister-chromatid exchange response in the presence of a mammalian metabolic activation system. Exposure of V79 Chinese hamster cells to 3,4-DCA caused a concentration-dependent increase in the incidence of spindle disturbances, predominantly of the initial c-mitotic type. The results indicate that 3,4-DCA might induce aneuploidy in mammalian cells by interaction with the mitotic apparatus. 相似文献