Bascomb N F, Yeung A T, Turner K J, Schmidt R R
J Bacteriol. 1981 Mar;145(3):1266-72. doi: 10.1128/jb.145.3.1266-1272.1981.
When ammonia was removed from Chlorella sorokiniana cells, which contain an ammonium-inducible nicotinamide adenine dinucleotide phosphate-specific glutamate dehydrogenase (NADP-GDH), the activity of this enzyme decayed with a half-life of approximately 8 min. By use of rocket immunoelectrophoresis, indirect immunoprecipitation, and indirect immunoadsorption (coupled with pulse-chase experiments with 35S-labeled sulfate), the rapid initial loss in activity was shown to be due to enzyme inactivation rather than degradation of NADP-GDH antigen. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of immunoprecipitates obtained with anti-NADP-GDH immunoglobulin G showed that enzyme inactivation is accompanied by the conversion of enzyme subunits (Mr = 59,000) to a protein with a molecular weight of 118,000. Because this protein was stable during boiling and in the presence of sodium dodecyl sulfate and high concentrations of mercaptoethanol or dithiothreitol, it was tentatively assumed to be a covalently linked dimer of enzyme subunits. Pulse-chase experiments showed that total NADP-GDH antigen was subject to rapid degradation (t 1/2 = 88 min) in induced cells, and the same degradation rate was maintained after removal of ammonia from induced cells.
当从小球藻细胞中去除氨时，该细胞含有一种铵诱导型烟酰胺腺嘌呤二核苷酸磷酸特异性谷氨酸脱氢酶（NADP-GDH），这种酶的活性以约8分钟的半衰期衰减。通过火箭免疫电泳、间接免疫沉淀和间接免疫吸附（结合用35S标记的硫酸盐进行的脉冲追踪实验），结果表明，酶活性最初的快速丧失是由于酶失活，而不是NADP-GDH抗原的降解。用抗NADP-GDH免疫球蛋白G获得的免疫沉淀物进行十二烷基硫酸钠-聚丙烯酰胺凝胶电泳显示，酶失活伴随着酶亚基（Mr = 59,000）转化为分子量为118,000的蛋白质。由于这种蛋白质在煮沸以及存在十二烷基硫酸钠和高浓度的巯基乙醇或二硫苏糖醇的情况下是稳定的，因此初步假定它是酶亚基的共价连接二聚体。脉冲追踪实验表明，在诱导细胞中，总NADP-GDH抗原会快速降解（t 1/2 = 88分钟），从诱导细胞中去除氨后，降解速率保持不变。
