Bellino F L, Hussa R O, Osawa Y
Steroids. 1978 Jul-Aug;32(1):37-44. doi: 10.1016/0039-128x(78)90097-1.
The stimulation of estrogen biosynthesis by N6,O2'--dibutyryl adenosine 3':5'-cyclic monophosphate and theophylline dbT) in cultures of the JAr line of choriocarcinoma cells was investigated by measuring the specific activity and kinetic constants of estrogen synthetase (aromatase) in the various subcellular fractions after differential centrifugation of homogenized cells in isotonic sucrose. The low speed (900xg) pellet, from cells grown with or without dbT and homogenized in isotonic sucrose, contains the majority of the aromatase activity and the highest aromatase specific activity. The aromatase specific activity in the homogenate of cells grown with dbT and in the various subcellular fractions is 4- to 10-fold higher than in cells grown without dbT. The Vmax of androstenedione (4-androstene-3,17-dione) aromatization in homogenates from dbT-stimulated cells (6.9 pmol estrogen/min per mg protein) is significantly increased over that measured in the absence of dbT (1.5 pmol estrogen/min per mg protein); the Km values, however, are not significantly different (average of 43.8nM in dbT-stimulated fractions; 53.2nM in control fractions). These results suggest that the increased aromatase specific activity in dbT-stimulated cells results from an increase in amount of active enzyme, rather than from an increase in affinity of the enzyme for its substrate.
通过在等渗蔗糖中对匀浆细胞进行差速离心后，测量各亚细胞组分中雌激素合成酶（芳香化酶）的比活性和动力学常数，研究了N6,O2'-二丁酰腺苷3':5'-环一磷酸和茶碱（dbT）对绒癌JAr细胞系培养物中雌激素生物合成的刺激作用。在等渗蔗糖中匀浆的、无论有无dbT培养的细胞，其低速（900xg）沉淀含有大部分芳香化酶活性和最高的芳香化酶比活性。用dbT培养的细胞匀浆及各亚细胞组分中的芳香化酶比活性比未用dbT培养的细胞高4至10倍。dbT刺激的细胞匀浆中雄烯二酮（4-雄烯-3,17-二酮）芳香化的Vmax（6.9 pmol雌激素/分钟·毫克蛋白）比未用dbT时测得的值（1.5 pmol雌激素/分钟·毫克蛋白）显著增加；然而，Km值没有显著差异（dbT刺激组分的平均值为43.8nM；对照组分的平均值为53.2nM）。这些结果表明，dbT刺激的细胞中芳香化酶比活性增加是由于活性酶量的增加，而不是由于酶对其底物亲和力的增加。
