Young D, Riggs M, Field J, Vojtek A, Broek D, Wigler M
Cold Spring Harbor Laboratory, NY 11724.
Proc Natl Acad Sci U S A. 1989 Oct;86(20):7989-93. doi: 10.1073/pnas.86.20.7989.
We cloned the adenylyl cyclase gene from the fission yeast Schizosaccharomyces pombe using low-stringency hybridization to the Saccharomyces cerevisiae adenylyl cyclase gene. The Sc. pombe gene encodes a 1692-amino acid-residue protein. The identity of this gene was confirmed by studies of its expression in Sa. cerevisiae. Expression of the carboxyl-terminal region of the Sc. pombe adenylyl cyclase protein will suppress a temperature-sensitive mutation in the Sa. cerevisiae adenylyl cyclase gene. Furthermore, Sa. cerevisiae that lack their endogenous adenylyl cyclase gene and express the carboxyl-terminal region of the Sc. pombe adenylyl cyclase protein have measurable adenylyl cyclase activity. The carboxyl-terminal region of this protein has strong homology with the catalytic domain of the Sa. cerevisiae adenylyl cyclase. Also, Sc. pombe adenylyl cyclase, like Sa. cerevisiae adenylyl cyclase, contains a tandemly repeated motif rich in leucine. Neither yeast protein is particularly homologous to the recently cloned Gs-responsive mammalian adenylyl cyclase [Krupinski, J., Coussen, F., Bakalyar, H. A., Tang, W.-J., Feinstein, P. G., Orth, K., Slaughter, C., Reed, R. R. & Gilman, A. G. (1989) Science 244, 1558-1564].
我们利用对酿酒酵母腺苷酸环化酶基因的低严谨度杂交,从裂殖酵母粟酒裂殖酵母中克隆了腺苷酸环化酶基因。粟酒裂殖酵母基因编码一种含有1692个氨基酸残基的蛋白质。通过对其在酿酒酵母中的表达研究证实了该基因的身份。粟酒裂殖酵母腺苷酸环化酶蛋白羧基末端区域的表达将抑制酿酒酵母腺苷酸环化酶基因中的温度敏感突变。此外,缺乏内源性腺苷酸环化酶基因并表达粟酒裂殖酵母腺苷酸环化酶蛋白羧基末端区域的酿酒酵母具有可测量的腺苷酸环化酶活性。该蛋白的羧基末端区域与酿酒酵母腺苷酸环化酶的催化结构域具有高度同源性。而且,粟酒裂殖酵母腺苷酸环化酶与酿酒酵母腺苷酸环化酶一样,含有一个富含亮氨酸的串联重复基序。这两种酵母蛋白与最近克隆的Gs反应性哺乳动物腺苷酸环化酶均无特别同源性[克鲁平斯基,J.,库森,F.,巴卡利亚尔,H. A.,唐,W.-J.,费恩斯坦,P. G.,奥思,K.,斯劳特,C.,里德,R. R. & 吉尔曼,A. G.(1989年)《科学》244, 1558 - 1564]。