Two cytoplasmic domains of mammalian adenylyl cyclase form a G(s alpha)- and forskolin-activated enzyme in vitro

被引:89
作者
Yan, SZ [1 ]
Hahn, D [1 ]
Huang, ZH [1 ]
Tang, WJ [1 ]
机构
[1] UNIV CHICAGO,DEPT PHARMACOL & PHYSIOL SCI,CHICAGO,IL 60637
关键词
D O I
10.1074/jbc.271.18.10941
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Mammalian adenylyl cyclases have two homologous cytoplasmic domains (C-1 and C-2). The first cytoplasmic domain of type I enzyme (IC1) and the second cytoplasmic domain of type II enzyme (IIC2-Delta 3, a construct in which 36 N-terminal amino acids of the C-2 region are deleted) were expressed and purified to homogeneity. Alone, each had no adenylyl cyclase activity; however, mixing of the two domains in vitro resulted in G(s alpha)- and forskolin activated enzyme activity. The turnover number for G(s alpha)- and forskolin-stimulated enzyme activity of the complex between IC1 and IIC2-Delta 3 was 8.2 s(-1). The concentration of IIC2-Delta 3 to achieve half-maximal activation of IC1 was 0.8 and 1.3 mu M when stimulated by forskolin and G(s alpha), respectively. The concentration of IIC2-Delta 3 needed to complex with IC1 was reduced 10-fold (0.08 mu M) when the enzyme was activated by both forskolin and G(s alpha), suggesting that G(s alpha) and forskolin increased the affinity of the two cytoplasmic domains for each other.
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页码:10941 / 10945
页数:5
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