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Induction of the enzyme Δ5-3-ketosteroid isomerase in Pseudomonas testosteroni was found to be strongly inhibited by reserpine and by the alkaloids of Vinca and Ergot. Morphine, colchicine and papaverine caused weaker inhibition whilst a series of other alkaloids were almost ineffective. Ergot alkaloids were inhibitory towards all steroids tested, androgens, oestrogens and progestagens, and a similar effect was shown with the other inducible enzymes, 3α- and 3β.17β-hydroxysteroid-dehydrogenase.
Experiments with cell-free protein synthesis indicate that reserpine inhibits the induction of messenger RNA.
Methylthio-β.ᴅ-galaktosid wird in E. coli K 12, sowie in den Mutanten ML 3 und ML 308 in vivo zu einem geringen Teil in einen Phosphorsäureester, wahrscheinlich das 6-Phosphat (TMG-P) umgewandelt. TMG-P wird von E. coli K 12 aufgenommen, wirkt jedoch nicht als Induktor des Lactose-Operons. Zellfreie Extrakte aus E. coli K 12 geben die gleiche Reaktion, wobei die in vitro-Reaktion durch anorganisches Phosphat und Phosphoenolpyruvat stimuliert wird.
The effect of NNMG on the template activities of different polynucleotides (polyuridylic acid, polycytidylic acid, polyadenylic acid and copolymer of adenylic and guanylic acid 5,5:1) and t-RNS was studied. The maximum inhibition of the messenger activity was found for poly-C, followed by poly-Α and poly-U. The acceptor activity of t-RNA was found to be inhibited by NNMG: maximum for proline, followed by serine, leucine, phenylalanine and lysine. The mechanism of these inhibitions was studied using NNMG radioactively labelled on the methyl group. Different amounts of radioactivity were found in the various polynucleotides and t-RNS.
Following treatment with the β-galactosidase inducer [methyl-3H] -thiogalactoside, an induceracceptor-complex was isolated from extracts of E. coli K 12 using DEAE cellulose chromatography. Enzymatic digestion with trypsin suggested that the inducer was bound to a protein component.
Specific radioactive peaks demonstrated acceptor activity in the inducible strains E. coli K 12 and ML 3, but different results were obtained using the non-inducible mutants ML 35, ML 308 and ML 309.
The potent inhibitor of TMG-induction, o-nitrophenylfucoside, reduced the radioactive acceptor peak and caused a similar inhibition of β-galactosidase synthesis, p-nitrophenylfucoside was ineffective.
Further evidence is presented for the in vitro formation of an inducer-acceptor-complex in cell free extracts of E. coli K 12.
The trifluoroacetylation of thymidine at room temperature was performed using trifluoroacetic acid phenylester in pyridine. A selective protection of the 5′-position was not possible: Even low molar quantities of the trifluoroacetylating agent gave rise to bis-trifluoroacetylation. The bis-trifluoroacetyl derivatives of thymidine and 5-bromo-deoxyuridine were purified by vacuum sublimation. The completely trifluoroacetylated deoxyribosides of uracil, 5-iodouracil and adenine underwent decomposition during sublimation.
The non-specific inhibition of the poly U directed polymerisation of phenylalanine through polyanions was studied. This inhibition was found to be in order as follows: dextransulfate, polyethylensulfate, heparine, ribosomal RNA and alginate. It was found that poly A, poly AP and poly AG cause a specific inhibition of the poly U directed synthesis of polyphenylalanine. Poly AG and poly AP, but not poly A were found to inhibit the poly C directed polymerisation of proline as well. The mechanism of these two types of inhibition caused by polyanions has been discussed.
It is possible to determine the anomeric configuration of nucleosides by a simple, spectrophotometric assay, using the nucleoside phosphorylase activity of cell-free extracts from E. coli. β-nucleo-sides are split, a-anomers remain unchanged. For a single estimation 20-40 µg of nucleoside are required. 6-Azauracil- and 8-azaguanine-β-ᴅ-riboside and some nucleoside phosphates are resistant, a fact, which is of interest in view of the specificity of nucleoside phosphorylases.
Steroid initiated enzyme induction (Δ5-Ketosteroid-Isomerase, 3α-Hydroxysteroid-Dehydrogenase, and 3β.17β-Hydroxysteroid-Dehydrogenase) in Pseudomonas testosteroni was investigated with respect to the kinetics of induction, operon control of the induced enzymes, and the relative strengths of various inducers. The induction process was followed indirectly by selective inhibition of different stages in the protein synthetic pathway. Comparisons between bacterial and mammalian steroid induction are discussed.
Bei der UV-Bestrahlung von Uracil-[5.6-3H] bilden sich je nach eingestrahlter Energie dimeres Uracil und Uracil-Wasseranlagerungsprodukt [5.6-Dihydro-6-hydroxyuracil] als radioaktive Photoprodukte. Während bei der Synthese des Wasseranlagerungsproduktes ein beträchtlicher sekundärer Isotopeneffekt wirksam wird, verändert sich die Radioaktivität des dimeren Uracils gegenüber der des Ausgangsuracils kaum.
Wird das Wasseranlagerungsprodukt durch Erwärmen zu Uracil zurückgewandelt, so dehydratisiert das Molekül ebenfalls unter Mitwirkung eines Isotopeneffektes. Wird das Uracildimere zu Uracil rückgewandelt, so beobachtet man keinen Isotopeneffekt.
Bei der Bestrahlung von Uracil in Tritium-haltigem Wasser werden nur sehr geringe Radioaktivitäten in die Photoprodukte eingebaut. Der Isotopeneffekt beträgt ca. 8. — Durch Synthese der Photoprodukte aus spezifisch an C-5 oder C-6 Tritium-markiertem Uracil bzw. durch Bromierung von 5.6-Tritium-markiertem Uracil bzw. dessen Photoprodukten zu den 5-Brom-Derivaten erhält man Hinweise, daß der Geschwindigkeits-bestimmende Schritt der Wasseraddition an C-6 des Uracils verläuft. Die inversen sekundären Isotopeneffekte betragen für Tritium an C-6 etwa 0,65, für Tritium an C-5 dagegen nur 0,95.