Mintzer K A, Field J
Department of Pharmacology, University of Pennsylvania School of Medicine, Philadelphia 19104, USA.
Cell Signal. 1999 Feb;11(2):127-35. doi: 10.1016/s0898-6568(98)00044-8.
Cdc25 and Ras are two proteins required for cAMP signalling in the budding yeast Saccharomyces cerevisiae. Cdc25 is the guanine nucleotide exchange protein that activates Ras. Ras, in turn, activates adenylyl cyclase. Cdc25 has a Src homology 3 (SH3) domain near the N-terminus and a catalytic domain in the C-terminal region. We find that a point mutation in the SH3 domain attenuates cAMP signalling in response to glucose feeding. Furthermore, we demonstrate, by using recombinant adenylyl cyclase and Cdc25, that the SH3 domain of Cdc25 can bind directly to adenylyl cyclase. Binding was specific, because the SH3 domain of Abp1p (actin-binding protein 1), which binds the 70,000 Mr subunit of adenylyl cyclase, CAP/Srv2, failed to bind adenylyl cyclase. A binding site for Cdc25-SH3 localised to the C-terminal catalytic region of adenylyl cyclase. Finally, pre-incubation with Ras enhanced the SH3-bound adenylyl cyclase activity. These studies suggest that a direct interaction between Cdc25 and adenylyl cyclase promotes efficient assembly of the adenylyl cyclase complex.
Cdc25和Ras是出芽酵母酿酒酵母中cAMP信号传导所需的两种蛋白质。Cdc25是激活Ras的鸟嘌呤核苷酸交换蛋白。反过来，Ras激活腺苷酸环化酶。Cdc25在N端附近有一个Src同源3（SH3）结构域，在C端区域有一个催化结构域。我们发现SH3结构域中的一个点突变会减弱对葡萄糖喂养的cAMP信号传导。此外，我们通过使用重组腺苷酸环化酶和Cdc25证明，Cdc25的SH3结构域可以直接与腺苷酸环化酶结合。这种结合是特异性的，因为与腺苷酸环化酶70,000 Mr亚基CAP/Srv2结合的Abp1p（肌动蛋白结合蛋白1）的SH3结构域未能与腺苷酸环化酶结合。Cdc25-SH3的结合位点定位于腺苷酸环化酶的C端催化区域。最后，与Ras预孵育增强了SH3结合的腺苷酸环化酶活性。这些研究表明，Cdc25与腺苷酸环化酶之间的直接相互作用促进了腺苷酸环化酶复合物的有效组装。
