Sun B, Jeyaseelan K, Chung M C, Tan T W, Chock P B, Teo T S
Department of Biochemistry, Faculty of Medicine, National University of Singapore.
Biochim Biophys Acta. 1997 Mar 20;1351(1-2):231-8. doi: 10.1016/s0167-4781(96)00209-6.
A cDNA clone encoding rabbit E2(32k) was obtained by library screening and PCR. The cDNA contains an open reading frame coding for 238 amino acids which shows an overall identity of 81% to human CDC34, the cell cycle-related ubiquitin-conjugating enzyme. A 50% homology to yeast CDC34 within the conserved core domain was also observed. Northern blot analysis indicated that three transcripts existed in all six rabbit tissues examined but their expression levels varied over a wide range. The putative cDNA coding region was highly expressed in Escherichia coli as a his-tagged protein which was purified to homogeneity. The ability of this expressed protein to form a thiolester bond with ubiquitin showed that it was functionally active. The ability of this protein to catalyze the conjugation of ubiquitin to histone H2A and H2B was also examined.
通过文库筛选和聚合酶链反应获得了编码兔E2(32k)的互补DNA（cDNA）克隆。该cDNA包含一个编码238个氨基酸的开放阅读框，与人类细胞周期相关的泛素结合酶CDC34的总体一致性为81%。在保守的核心结构域内也观察到与酵母CDC34有50%的同源性。Northern印迹分析表明，在所检测的所有六种兔组织中存在三种转录本，但它们的表达水平在很宽的范围内变化。推定的cDNA编码区在大肠杆菌中作为带有组氨酸标签的蛋白质高表达，并被纯化至同质。这种表达蛋白与泛素形成硫酯键的能力表明它具有功能活性。还检测了该蛋白催化泛素与组蛋白H2A和H2B结合的能力。
