Takio K, Smith S B, Krebs E G, Walsh K A, Titani K
Biochemistry. 1984 Aug 28;23(18):4200-6. doi: 10.1021/bi00313a029.
Evidence is presented that establishes the amino acid sequence of the regulatory subunit of type II cAMP-dependent protein kinase from bovine cardiac muscle. Complementary sets of overlapping peptides were generated primarily by tryptic digestion and by chemical cleavage at methionyl residues. The analysis was augmented by chemical cleavage at a single tryptophanyl residue and at three of the four aspartyl-proline bonds. Several large fragments generated by limited proteolysis contributed to the proof of structure. The subunit is a single chain of 400 residues corresponding to a molecular weight of 45 004. An amino-terminal segment of about 100 residues is believed to include the region responsible for oligomeric association. The remainder of the molecule consists of two tandem homologous domains, each of which is thought to bind a single molecule of cAMP. Comparison of the three domains with corresponding regions of the type I isozyme, of the Escherichia coli catabolite gene activator protein, and of cGMP-dependent protein kinase indicates extensive regions of homology and as much as 50% identity with the sequence of an internal segment of the type I isozyme.
本文提供的证据确定了牛心肌中II型cAMP依赖性蛋白激酶调节亚基的氨基酸序列。互补的重叠肽组主要通过胰蛋白酶消化以及在甲硫氨酰残基处进行化学裂解产生。通过在单个色氨酰残基处以及四个天冬氨酰-脯氨酸键中的三个处进行化学裂解，增强了分析效果。有限蛋白酶解产生的几个大片段有助于结构的证明。该亚基是一条由400个残基组成的单链，分子量为45004。约100个残基的氨基末端片段被认为包含负责寡聚体缔合的区域。分子的其余部分由两个串联的同源结构域组成，每个结构域被认为结合一分子cAMP。将这三个结构域与I型同工酶、大肠杆菌分解代谢基因激活蛋白以及cGMP依赖性蛋白激酶的相应区域进行比较，结果表明存在广泛的同源区域，并且与I型同工酶内部片段的序列有高达50%的同一性。
