Zhai L, Graves P R, Robinson L C, Italiano M, Culbertson M R, Rowles J, Cobb M H, DePaoli-Roach A A, Roach P J
Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis 46202-5122, USA.
J Biol Chem. 1995 May 26;270(21):12717-24. doi: 10.1074/jbc.270.21.12717.
Casein kinase I, one of the first protein kinases identified biochemically, is known to exist in multiple isoforms in mammals. Using a partial cDNA fragment corresponding to an isoform termed CK1 gamma, three full-length rat testis cDNAs were cloned that defined three separate members of this subfamily. The isoforms, designated CK1 gamma 1, CK1 gamma 2, and CK1 gamma 3, have predicted molecular masses of 43,000, 45,500, and 49,700. CK1 gamma 3 may also exist in an alternatively spliced form. The proteins are more than 90% identical to each other within the protein kinase domain but only 51-59% identical to other casein kinase I isoforms within this region. Messages for CK1 gamma 1 (2 kilobases (kb)), CK1 gamma 2 (1.5 and 2.4 kb), and CK1 gamma 3 (2.8 kb) were detected by Northern hybridization of testis RNA. Message for CK1 gamma 3 was also observed in brain, heart, kidney, lung, liver, and muscle whereas CK1 gamma 1 and CK1 gamma 2 messages were restricted to testis. All three CK1 gamma isoforms were expressed as active enzymes in Escherichia coli and partially purified. The enzymes phosphorylated typical in vitro casein kinase I substrates such as casein, phosvitin, and a synthetic peptide, D4. Phosphorylation of the D4 peptide was activated by heparin whereas phosphorylation of the protein substrates was inhibited. The known casein kinase I inhibitor CK1-7 also inhibited the CK1 gamma s although less effectively than the CK1 alpha or CK1 delta isoforms. All three CK1 gamma s underwent autophosphorylation when incubated with ATP and Mg2+. The YCK1 and YCK2 genes in Saccharomyces cerevisiae encode casein kinase I homologs, defects in which lead to aberrant morphology and growth arrest. Expression of mammalian CK1 gamma 1 or CK1 gamma 3 restored growth and normal morphology to a yeast mutant carrying a disruption of YCK1 and a temperature-sensitive allele of YCK2, suggesting overlap of function between the yeast Yck proteins and these CK1 isoforms.
酪蛋白激酶I是最早通过生物化学方法鉴定出的蛋白激酶之一，已知在哺乳动物中以多种同工型存在。利用对应于一种称为CK1γ同工型的部分cDNA片段，克隆了三个全长大鼠睾丸cDNA，它们定义了该亚家族的三个不同成员。这些同工型分别命名为CK1γ1, CK1γ2和CK1γ3，预测分子量分别为43,000、45,500和49,700。CK1γ3也可能以可变剪接形式存在。这些蛋白在蛋白激酶结构域内彼此的同源性超过90%，但在该区域内与其他酪蛋白激酶I同工型的同源性仅为51 - 59%。通过睾丸RNA的Northern杂交检测到CK1γ1（2千碱基（kb））、CK1γ2（1.5和2.4 kb）和CK1γ3（2.8 kb）的信使RNA。在脑、心脏、肾脏、肺、肝脏和肌肉中也观察到CK1γ3的信使RNA，而CK1γ1和CK1γ2的信使RNA仅限于睾丸。所有三种CK1γ同工型在大肠杆菌中均表达为活性酶并进行了部分纯化。这些酶可磷酸化典型的体外酪蛋白激酶I底物，如酪蛋白、卵黄高磷蛋白和一种合成肽D4。D4肽的磷酸化被肝素激活，而蛋白底物的磷酸化被抑制。已知的酪蛋白激酶I抑制剂CK1 - 7也抑制CK1γ同工型，尽管其效果不如CK1α或CK1δ同工型。当与ATP和Mg2 +一起孵育时，所有三种CK1γ同工型都进行了自身磷酸化。酿酒酵母中的YCK1和YCK2基因编码酪蛋白激酶I同源物，其缺陷会导致异常形态和生长停滞。哺乳动物CK1γ1或CK1γ3的表达恢复了携带YCK1破坏和YCK2温度敏感等位基因的酵母突变体的生长和正常形态，表明酵母Yck蛋白与这些CK1同工型之间存在功能重叠。
