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用电镜细胞化学方法研究了大白鼠肝细胞溶酶体膜的中性pHp—对硝基苯磷酸酶(p-nltrophenyl phosphatase)(p-NPPase)的活性。切片用野田等人报告的原法反应时,溶酶体膜和基质均有酶的活性。加入酸性磷酸酶(AcPase)的抑制剂氟化钠(NaF)后,基质的酶活性被抑制,膜上的则不受影响。相反,当孵育液中含有4,4′-diisothiocyanostilbene-2.2′-disulfonic acid,disodium salt(DIDS)或N—乙基顺丁稀二酰亚胺(N-ethylmaleimide)(NEM)时,溶酶体膜上的酶反应被抑制而基质反应不受影响。如将氟化钠或DIDS或氟化钠和N—乙基顺丁烯二酰亚胺同时加到孵育液中,则膜和基质的反应都消失。这些结果表明溶酶体膜上中性范围的p—NPPase活性可能代表了H~+—ATPase活性。
The activity of neutral pHp-p-nltrophenyl phosphatase (p-NPPase) in rat liver cell lysosome membranes was studied by electron microscopy cytochemistry. Slice with Noda et al. Reported the original method reaction, the lysosomal membrane and matrix enzyme activity. After adding sodium fluoride (NaF), an inhibitor of acid phosphatase (AcPase), the enzyme activity of the matrix was inhibited and the membrane was unaffected. In contrast, when the incubation solution contained 4,4’-diisothiocyanostilbene-2.2’-disulfonic acid, disodium salt (DIDS) or N-ethylmaleimide (NEM) The enzyme reaction on the membrane is inhibited and the matrix reaction is unaffected. If sodium fluoride or DIDS or sodium fluoride and N-ethyl maleimide are simultaneously added to the incubation solution, the reaction between the membrane and the substrate disappears. These results indicate that the neutral range of p-NPPase activity on lysosomal membranes may represent H ~ + -ATPase activity.