Construction of a soluble adenylyl cyclase activated by Gs alpha and forskolin.
Construction of a soluble adenylyl cyclase activated by Gs alpha and forskolin.
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构建由 Gs α 和毛喉素激活的可溶性腺苷酸环化酶。
DOI：
10.1126/science.7792604
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发表时间：
1995
期刊：
影响因子：
--
通讯作者：
Gilman,AG
中科院分区：
文献类型：
--
作者：
Tang,WJ;Gilman,AG
通过连接哺乳动物I型和II型酶的胞质结构域的部分来构建可溶性腺苷酸环化酶。可溶性酶受到毛喉素和异源三聚鸟嘌呤核苷酸结合蛋白（G 蛋白）Gs(Gsα) 的 α 亚基的刺激。该构建体的表达补充了缺乏腺苷酸环化酶活性的大肠杆菌菌株中的分解代谢缺陷。大约 60 千道尔顿的活性酶在大肠杆菌的细胞质部分中积累。大肠菌的产量活性超过每分钟每毫克蛋白质 1 纳摩尔。因此，哺乳动物腺苷酸环化酶的两组跨膜螺旋对于酶的催化或最具特征性的调节活性不是必需的。该系统可用于 G 蛋白调节的腺苷酸环化酶的遗传和生化分析。
A soluble adenylyl cyclase was constructed by linkage of portions of the cytosolic domains of the mammalian type I and type II enzymes. The soluble enzyme was stimulated by both forskolin and the α subunit of the heterotrimeric guanine nucleotide-binding protein (G protein) Gs(Gsα). Expression of the construct complemented the catabolic defect in a strain ofEscherichia colithat is deficient in adenylyl cyclase activity. The active, approximately 60-kilodalton enzyme accumulated in the cytoplasmic fraction ofE. colito yield activities in excess of 1 nanomole per minute per milligram of protein. The two sets of transmembrane helices of mammalian adenylyl cyclases are thus not necessary for the catalytic or the most characteristic regulatory activities of the enzyme. This system may be useful for both genetic and biochemical analysis of G protein-regulated adenylyl cyclases.