Studies on Nitro-Reducing Enzymes of Liver
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概要
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1. Seventy per cent of p-nitrophenol injected intraperitoneally to the rat was conjugatingly detoxicated at the hydroxyl group, while 30 per cent was reduced to p-aminophenol and excreted in the urine. 2. Foreign sulfate ion is utilized for the esterification of p-nitrophenol, after possible "activation" in the liver. 3. The capacity of the liver to esterify phenol with sulfate increased in some "adaptive" manner. 4. Several investigations of nitro-reducing capacity of the liver were performed. a) The liver, kidneys, heart, muscle, lung, testicles, spleen, uterus, vagina, and ovary of rats were homogenized and examined, and the former seven were found to be able to reduce p-nitrophenol in the above order. b) The reduction of the nitro group by the liver is inhibited by cyanide, monoiodoacetic acid, sodium azide, and thiourea, suggesting the indispensable participation of heavy metals in the reduction mechanism. c) Dialysis removed the reducing capacity of the liver, which is restored by the addition of a boiled juice of the liver, succinate, glucose plus DPN, and alcohol plus DPN. d) Acetone precipitation also removed the capacity, which was restored by the addition of the boiled juice, glucose and alcohol with DPN. e) The acetone fractionation of the liver homogenate affords a fraction which possesses nitro-reducing capacity only when coupled with alcohol dehydrogenase system (alcohol plus alcohol dehydrogenase plus DPN). The alcohol dehydrogenase system failed to reduce p-nitrophenol in the absence of the nitroreductase fraction of the liver. f) The difference in the order of the distribution between the nitro-reducing capacity and succinic dehydrogenase activity was shown and the existence of an enzyme was discussed which plays an indispensable role directly in the reduction of the aromatic nitro group in the mammalia. 5. Properties of nitroreductase enzyme of the liver has been investigated which is able to transfer the hydrogen atoms or electrons from reduced pyridine nucleotide to aromatic nitro compounds forming amines. Co factors and hydrogen carriers of the reducing reaction has been also studied and the roles of metal ions and flavine nucleotides has been established. 6. Swine liver nitroreductase enzyme was separated into two components, one acting upon nitro group forming nitroso, and another reducing nitroso to hydroxylamino or to amino, by means of the ion exchanger studies, and the name "nitroreductase" and "nitrosoreductase" were proposed for the two enzymes. The former enzymes has been suggested to catalyze the rate limiting step of the nitroreducing system of the liver. 7. Two nitroreductase enzymes of the liver which are responsible for the reductions of nitro and nitroso groups respectively, were isolated separately and partially purified column chromatographically with diethylaminoethyl-cellulose. 8. Several properties of the two enzymes have been studied: Both enzymes are active at pH 7.8. DPNH is about ten times more active than TPNH as the hydrogen donor for both enzyme reactions. Nitroreductase is inhibited by oxygen, whereas nitrosoreductase is quite insensitive to oxygen. Both of the enzyme actions are inhibited by several metal-chelating agents, suggesting that they are both metalloenzymes. The enzymes are considered to have the active sulfhydryl groups from the inhibitory studies. 9. Nitroreductase was found to be flavoenzyme which specifically utilize flavinadenine dinucleotide as the hydrogen carrier, whereas the flavine nucleotides are considered not to be necessary for the reduction of nitrosophenol by the action of nitrosoreductase.
- 大阪府立大学の論文
- 1961-03-31