Wang S L, Rice S A, Serra M T, Gross B
Drug Metab Dispos. 1986 Jul-Aug;14(4):392-8.
Enzymes responsible for the defluorination of methoxyflurane (MOF) and fluoroacetate (FAc) were separated and purified from rat liver cytosol. Both hepatic cytosolic enzymes with defluorination activity were labile and addition of 2-mercaptoethanol had little effect on the stability of these enzymes. Glutathione S-transferase (GT) activity of the same cytosolic fractions was stable for at least 11 days. Separation of defluorination and GT enzymatic activities on DEAE-Sephadex A-50 and reduced glutathione-affinity columns revealed that the defluorinations of MOF and FAc were primarily catalyzed by anionic proteins which also exhibited GT activity. Further identification by two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed protein bands with pl values of approximately 6.5 and 6.9 and molecular weights of approximately 20,000. However, other proteins that exhibited no GT activity also defluorinated MOF and FAc, but accounted for only 10% of the total defluorination activity present in anionic proteins. Results from a separate purification experiment using a CM-cellulose column also indicated that the enzymes responsible for defluorination coeluted with cationic GTs. Collectively, these cationic enzymes were responsible for about 20% of the recovered cytosolic defluorination activities. The results suggest that the cytosolic defluorinations of both MOF and FAc are primarily the result of a dehalogenation reaction catalyzed by one or more species of rat liver cytosolic GTs.
负责甲氧氟烷（MOF）和氟乙酸盐（FAc）脱氟的酶从大鼠肝细胞溶胶中分离并纯化。两种具有脱氟活性的肝细胞溶胶酶都不稳定，添加2-巯基乙醇对这些酶的稳定性影响很小。相同细胞溶胶组分的谷胱甘肽S-转移酶（GT）活性至少稳定11天。在DEAE-葡聚糖A-50和还原型谷胱甘肽亲和柱上分离脱氟和GT酶活性表明，MOF和FAc的脱氟主要由也表现出GT活性的阴离子蛋白催化。通过二维十二烷基硫酸钠-聚丙烯酰胺凝胶电泳进一步鉴定显示，蛋白带的等电点约为6.5和6.9，分子量约为20,000。然而，其他不表现出GT活性的蛋白也能使MOF和FAc脱氟，但仅占阴离子蛋白中总脱氟活性的10%。使用CM-纤维素柱的单独纯化实验结果也表明，负责脱氟的酶与阳离子GTs共洗脱。总的来说，这些阳离子酶约占回收的细胞溶胶脱氟活性的20%。结果表明，MOF和FAc的细胞溶胶脱氟主要是由一种或多种大鼠肝细胞溶胶GTs催化的脱卤反应的结果。
