Uno I, Oshima T, Ishikawa T
Institute of Applied Microbiology, University of Tokyo, Japan.
Exp Cell Res. 1988 Jun;176(2):360-5. doi: 10.1016/0014-4827(88)90337-0.
The subcellular distribution of the regulatory subunit of cAMP-dependent protein kinase in Saccharomyces cerevisiae cells was determined by subcellular fractionation and indirect immunofluorescence microscopy using the bcy1 mutant deficient in the regulatory subunit as control. The regulatory subunit of cAMP-dependent protein kinase showing cAMP-binding activity was identified as a single protein of 50 kDa by photoaffinity labeling and immunoblotting. The regulatory subunit was concentrated in a nuclear fraction in addition to a cytoplasmic fraction. By comparison of the regulatory subunit distribution with the DNA localization, the area detected by the indirect immunofluorescence was identified as the nucleus.
利用缺乏调节亚基的bcy1突变体作为对照，通过亚细胞分级分离和间接免疫荧光显微镜技术，确定了酿酒酵母细胞中cAMP依赖性蛋白激酶调节亚基的亚细胞分布。通过光亲和标记和免疫印迹法，显示具有cAMP结合活性的cAMP依赖性蛋白激酶调节亚基被鉴定为一种50 kDa的单一蛋白质。调节亚基除了存在于细胞质部分外，还集中在细胞核部分。通过将调节亚基的分布与DNA定位进行比较，间接免疫荧光检测到的区域被确定为细胞核。
