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The human class I alcohol dehydrogenase isozyme beta 2 beta 2 was used as the antigen to raise monoclonal antibodies. Altogether seven lines of hybridomas secreting monoclonal antibodies were obtained. None of the antibodies was isozyme specific and all of them exhibited a similar affinity against all isozymes of the human class I ADH. Five out of the seven monoclonal antibodies had no effect on beta 2 beta 2 activity. Antibody G3 acted as a non-competitive inhibitor with a KI of 3 micrograms/ml at pH 7.5. Increasing pH was effective in reducing the level of inhibition. On the other hand, antibody 1D4 exhibited a pH-dependent activation of ADH activity. In the presence of this antibody, the pH optimum of beta 2 beta 2 was shifted from 9 to 8.5 and total activity was increased by 70% at this optimal pH. Kinetic analysis indicates that 1D4 probably acts as a non-competitive activator and may exert its action by interacting with the coenzyme binding site. 相似文献
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Moore MR Spence JB Kiningham KK Dillon JL 《The Journal of steroid biochemistry and molecular biology》2006,98(4-5):218-227
Previously, we have shown that progestins both stimulate proliferation of the progesterone receptor (PR)-rich human breast cancer cell line T47D and protect from cell death, in charcoal-stripped serum-containing medium. To lessen the variability inherent in different preparations of serum, we decided to further characterize progestin inhibition of cell death using serum starvation to kill the cells, and find that progestins protect from serum-starvation-induced apoptosis in T47D cells. This effect exhibits specificity for progestins and is inhibited by the antiprogestin RU486. While progestin inhibits cell death in a dose–responsive manner at physiological concentrations, estradiol-17β surprisingly does not inhibit cell death at any concentration from 0.001 nM to 1 μM. Progestin inhibition of cell death also occurs in at least two other human breast cancer cell lines, one with an intermediate level of PR, MCF-7 cells, and, surprisingly, one with no detectable level of PR, MDA-MB-231 cells. Further, we have found progestin inhibition of cell death caused by the breast cancer chemotherapeutic agents doxorubicin and 5-fluorouracil. These data are consistent with the building body of evidence that progestins are not the benign hormones for breast cancer they have been so long thought to be, but may be harmful both for undiagnosed cases and those undergoing treatment. 相似文献
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S Pontremoli E Melloni F Salamino B Sparatore M Michetti O Sacco B L Horecker 《Biochemical and biophysical research communications》1986,140(3):1121-1126
Exposure of human neutrophils to low concentrations of phorbol myristate acetate (PMA) results, after a brief lag, in the production of superoxide anion and the phosphorylation of membrane proteins. Evidence that these responses are linked has now been obtained using a monoclonal antibody directed against an undefined macrophage surface antigen. The addition of this antibody, which recognizes a 90 kDa neutrophil membrane protein, caused dose-dependent delays in the onset of both phosphorylation of neutrophil membrane proteins and in the appearance of superoxide anion, following addition of PMA to the cell suspensions. For each response the lag period increased with increasing concentrations of antibody, but the onset of phosphorylation always preceded by a few minutes the initial appearance of superoxide anion. 相似文献
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Selective inhibition of human lung cancer cell growth by peptides derived from retinoblastoma protein 总被引:1,自引:0,他引:1
Peptides containing retinoblastoma protein (RB) fragment 649-654 (LFYKKV) were tested for their ability to block the proliferation of RB-negative and RB-positive human non-small cell lung cancer (NSCLC) cells. These peptides potently restrained the growth of both types of tumor cells, as measured by metabolic (MTT) and cellular viability (trypan blue exclusion) assays. As such, and remarkably, the peptides were able to overcome the resistance of RB-positive cells usually observed with RB gene or protein replacement therapy. Compared to the overall performance of conventional chemotherapy tested in parallel, the peptides were more cytotoxic against RB-negative neoplastic cells and equipotent toward RB-positive tumor cells, yet less toxic toward normal human cells. Thus, these new molecules hold great promise to evolve into an efficient therapy for human lung cancer, a common malignancy still defying treatment and holding a poor prognosis, as well as for other human neoplasias. 相似文献
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Activation and inhibition of transcription by supercoiling 总被引:18,自引:0,他引:18
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Initial rate kinetics of dextran synthesis by dextransucrase (sucrose:1,6-alpha-D-glucan-6-alpha-D-glucosyltransferase, EC 2.4.1.5) from Leuconostoc mesenteroides NRRL B-512F showed that below 1 mM, Ca2+ activated the enzyme by increasing Vmax and decreasing the Km for sucrose. Above 1 mM, Ca2+ was a weak competitive inhibitor (Ki = 59 mM). Although it was an activator at low concentration, Ca2+ was not required for dextran synthesis, either of main chain or branch linkages. Neither was it required for sucrose hydrolysis, acceptor reactions, or enzyme renaturation after SDS-polyacrylamide gel electrophoresis. A model for dextran synthesis is proposed in which dextransucrase has two Ca2+ sites, one activating and one inhibitory. Ca2+ at the inhibitory site prevents the binding of sucrose. 相似文献
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Enhanced TRAIL sensitivity by E1A expression in human cancer and normal cell lines: inhibition by adenovirus E1B19K and E3 proteins 总被引:2,自引:0,他引:2
Hu B Zhu H Qiu S Su Y Ling W Xiao W Qi Y 《Biochemical and biophysical research communications》2004,325(4):1153-1162
TNF-related apoptosis-inducing ligand (TRAIL) is a member of the TNF superfamily of cytokines that induces apoptosis in a variety of cancer cells, but not in normal cells. However, more and more tumor cells remain resistant to TRAIL, which limited its application for cancer therapy. Expression of the adenovirus serotype 5 (Ad5) E1A sensitizes tumor cells to apoptosis by TNF-alpha, Fas-ligand, and TRAIL. Here we asked whether E1A overcomes this resistance and enhances TRAIL-induced apoptosis in the tumor cells. Our results revealed that the tumor cell lines, HeLa and HepG2, with infection by Ad-E1A, were highly sensitive to TRAIL-induced apoptosis. Importantly, we found that in normal primary human lung fibroblast cells (HLF) TRAIL is capable of inducing apoptosis in combination with E1A as efficiently as in some tumor cell lines. The adenovirus type 5 encoding proteins, E1B19K and E3 gene products, have been shown to inhibit E1A and TRAIL-induced apoptosis of HLF cells by using the recombinant adenovirus AdDeltaE1B55K, with mutation of E1B55K, containing E1B19K and complete E3 region. Further results demonstrated that the expression of DR5 and TRAIL was down-regulated in the AdDeltaE1B55K co-infected HLF cells. These findings suggest that TRAIL may play an important role in limiting virus infections and the ability of adenovirus to inhibit killing may prolong acute and persistent infections. The results from this study have also suggested the possibility that the combination of E1A with TRAIL could be used in the treatment of human malignancy, or in the selection of the optimal adenovirus mutant as effective delivering vector for cancer therapy. 相似文献
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N Terakawa H Ikegami I Shimizu T Aono O Tanizawa K Matsumoto 《Journal of steroid biochemistry》1987,28(5):571-574
Since we recently found that danazol, an isoxazol derivative of ethinyltestosterone, has a growth-inhibitory effect on human endometrial cancer cells in primary culture, the effects of danazol on a human endometrial cancer cell line (IK-90 cells), which contains estrogen-independent progesterone receptors (PR), were investigated in the present study. The addition of danazol (1 nM-1 microM) in culture medium caused a decrease in the growth of IK-90 cells in a dose-dependent manner. Competitive binding studies showed that danazol effectively binds to PR in IK-90 cells, and the binding affinity for PR was estimated to be 6.0% of that of R5020. The addition of 1 microM danazol in culture medium resulted in a rapid and significant increase in nuclear PR with a concomitant decrease in cytoplasmic PR in the cells. These findings suggest that danazol has a growth-inhibitory effect on human endometrial adenocarcinoma cells directly through PR system in the cells. 相似文献
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Two-dimensional liquid separations-mass mapping of proteins from human cancer cell lysates 总被引:7,自引:0,他引:7
Lubman DM Kachman MT Wang H Gong S Yan F Hamler RL O'Neil KA Zhu K Buchanan NS Barder TJ 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》2002,782(1-2):183-196