Mendoza-Hernández G, Libreros-Minotta C A, Rendón J L
Departamento de Bioquímica, Facultad de Medicina, Universidad Nacional Autónoma de México, D.F., México.
Comp Biochem Physiol B Biochem Mol Biol. 1996 Oct;115(2):273-9. doi: 10.1016/0305-0491(96)00115-0.
Functional 3 beta-hydroxysteroid dehydrogenase coupled with isomerase (3 beta-HSD) was extracted from dog pancreatic mitochondria by treatment with the zwitterionic detergent CHAPSO. Increasing concentrations of this detergent led to a progressive and simultaneous solubilization of the pregnene (C-21) and androstene (C-19) dehydrogenase activities. Optimal solubilization of both C-21 and C-19 3 beta-HSD activities was achieved at a detergent/protein ratio of 0.6 (w/w). One hundred thirty percent of the initial particulate enzyme activities were recovered in the 105,000 g supernatant fluid with a 2.5-fold increase in the enzymatic specific activities. The C-21/C-19 activity ratios were 1.3 for mitochondria and 1.39 for the solubilized preparation. The apparent Km values for steroid substrates were unchanged after solubilization. Treatment of the mitochondrial suspension with sodium deoxycholate, CTAB, Lubrol XW, Brij 58, Emulgen 913 and Triton X-100 markedly decreased the 3 beta-HSD activities as a function of the detergent concentration and failed in to achieve solubilization.
通过用两性离子去污剂CHAPSO处理，从犬胰腺线粒体中提取了功能性3β-羟基类固醇脱氢酶与异构酶（3β-HSD）。该去污剂浓度的增加导致孕烯（C-21）和雄烯（C-19）脱氢酶活性逐渐同时溶解。在去污剂/蛋白质比为0.6（w/w）时，C-21和C-19 3β-HSD活性均实现了最佳溶解。在105,000 g的上清液中回收了130%的初始颗粒酶活性，酶比活性增加了2.5倍。线粒体的C-21/C-19活性比为1.3，溶解制剂的该比值为1.39。溶解后，类固醇底物的表观Km值未发生变化。用脱氧胆酸钠、十六烷基三甲基溴化铵、Lubrol XW、Brij 58、乳化剂913和Triton X-100处理线粒体悬液，3β-HSD活性随去污剂浓度显著降低，且未能实现溶解。
