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By simple substitution of an N-acyl group for the anthraniloyl(o-aminobenzoyl) group, chromogenic p-nitroanilide substrates are converted into highly sensitive fluorogenic substrates of proteases. The fluorescence of the anthraniloyl group is completely quenched by the p-nitroanilide moiety in the intact substrates and is released during their enzymatic hydrolysis. The approach is exemplified by the synthesis of anthraniloyl-Phe p-nitroanilide, anthraniloyl-Lys p-nitroanilide, and anthraniloyl-Gly-Gly-Phe p-nitroanilide as substrates for chymotrypsin, trypsin, and alkaline mesentericopeptidase, respectively. The kinetic parameters of these substrates are slightly better than those of similar derivatives bearing other acyl groups, suggesting that the enhanced sensitivity is completely due to the method of measurement. Since the conversion does not affect the chromogenic properties of the substrates, the same compounds can be used as usual p-nitroanilide substrates as well. 相似文献
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A Banerjee M A Jordan R F Luduena 《Biochemical and biophysical research communications》1985,128(2):506-512
Incubation of phosphocellulose-purified tubulin with GSH at 30 degrees C results in an inhibition of colchicine binding activity. GSSG has a protective effect against the GSH-induced loss of colchicine-binding. Incubation of tubulin with GSH at 30 degrees C results in the formation of abnormal tubulin polymers which are insensitive to cold. Such aggregation is insensitive to antimicrotubular drugs. Aggregation is inhibited by GSSG but not by DTT or mercaptoethanol. GSH-induced aggregation is very sensitive to the ionic strength of the assembly medium; both the aggregation and colchicine binding inhibition induced by GSH are inhibited at higher ionic strength. These results indicate a very complex interaction of GSH with tubulin. 相似文献
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The present investigation was made to evaluate biologically relevant quinones as possible catalysts in the generation of hydroxyl radicals from hydrogen peroxide and superoxide radicals. ESR spectra demonstrated that menadione, plastoquinone, and ubiquinone derivatives could all be reduced to their semiquinone forms by electron transfer from superoxide radicals. Reductive homolytic cleavage of H2O2 was observed to be dependent upon the presence of semiquinones in the reaction medium. Our data strongly support the concept that quinones normally involved in physiological processes may also play a role as catalysts of the Haber-Weiss reaction in the biological generation of hydroxyl radicals. 相似文献