Clegg C H, Cadd G G, McKnight G S
Department of Pharmacology, University of Washington, Seattle 98195.
Proc Natl Acad Sci U S A. 1988 Jun;85(11):3703-7. doi: 10.1073/pnas.85.11.3703.
An isoform (RI beta) of the regulatory type I subunit gene of cAMP-dependent protein kinase (EC 2.7.1.37) has been characterized in mouse. The open reading frame of the RI beta cDNA is 72% identical in nucleotide sequence with the previously cloned RI gene, now referred to as RI alpha. Both genes code for a protein of 380 amino acids and their proteins are 82% identical in amino acid sequence. Sequence similarity is highest in the regions that form the pseudosubstrate-binding site of the catalytic subunit and the two cAMP binding domains. The amino-terminal portion shows the greatest dissimilarity, suggesting that the isoforms may differ in their dimerization properties or interaction with other proteins. In contrast to RI alpha, which is constitutively expressed in all tissues, RI beta is expressed in a highly tissue-specific manner. Brain and spinal cord contained significant levels of RI beta mRNA, testis RNA gave a detectable signal, and all other tissues tested were negative. Expression of a RI beta cDNA in NIH 3T3 cells resulted in the appearance of a RI subunit protein that migrated more slowly than RI alpha after NaDodSO4/PAGE. The native form of RI beta in brain could also be distinguished from RI alpha by its abnormal migration on NaDodSO4/PAGE. RI beta protein produced in 3T3 cells was shown to be functional by its ability to form a cAMP-dependent holoenzyme with the catalytic subunit.
已在小鼠中鉴定出环磷酸腺苷依赖性蛋白激酶（EC 2.7.1.37）调节性I型亚基基因的一种同工型（RIβ）。RIβ cDNA的开放阅读框在核苷酸序列上与先前克隆的RI基因（现称为RIα）有72%的同一性。这两个基因都编码一种由380个氨基酸组成的蛋白质，它们的蛋白质在氨基酸序列上有82%的同一性。在形成催化亚基的假底物结合位点和两个环磷酸腺苷结合结构域的区域，序列相似性最高。氨基末端部分差异最大，这表明同工型在其二聚化特性或与其他蛋白质的相互作用方面可能有所不同。与在所有组织中组成性表达的RIα不同，RIβ以高度组织特异性的方式表达。脑和脊髓含有大量的RIβ mRNA，睾丸RNA给出了可检测的信号，而所有其他测试组织均为阴性。在NIH 3T3细胞中表达RIβ cDNA导致出现一种RI亚基蛋白，在十二烷基硫酸钠/聚丙烯酰胺凝胶电泳（NaDodSO4/PAGE）后，其迁移速度比RIα慢。脑中RIβ的天然形式在NaDodSO4/PAGE上的异常迁移也可与RIα区分开来。在3T3细胞中产生的RIβ蛋白通过其与催化亚基形成环磷酸腺苷依赖性全酶的能力被证明具有功能。
