Walter U, Greengard P
J Cyclic Nucleotide Res. 1978 Dec;4(6):437-44.
Several methods were compared for estimating the amount of regulatory subunit of an 800-fold purified Type II cAMP-dependent protein kinase from bovine heart. These methods included a reversable binding assay using either cAMP, or 8-N3-[32P]cAMP, photoaffinity labeling with 8-N3-[32P]cAMP, and autophosphorylation of the regulatory subunit of the enzyme. Although the regulatory subunit had a slightly lower affinity for 8-N3-cAMP than for cAMP, the total amount of regulatory subunit could be determined by each of the procedures examined. The results indicate that the photoaffinity analog 8-N3-[32P]cAMP is able to label quantitatively all cAMP-binding sites of the regulatory subunit of this cAMP-dependent protein kinase.
为了估算从牛心脏中提取的纯化800倍的II型环磷酸腺苷（cAMP）依赖性蛋白激酶调节亚基的含量，对几种方法进行了比较。这些方法包括使用cAMP或8-N3-[32P]cAMP的可逆结合测定、用8-N3-[32P]cAMP进行光亲和标记以及该酶调节亚基的自磷酸化。尽管调节亚基对8-N3-cAMP的亲和力略低于对cAMP的亲和力，但通过所研究的每种方法都可以确定调节亚基的总量。结果表明，光亲和类似物8-N3-[32P]cAMP能够定量标记这种cAMP依赖性蛋白激酶调节亚基的所有cAMP结合位点。
