Yang Liuqing, Yang Jenny, Huang Youliang, Liu Zhi Ren
Department of Biology, Georgia State University, Atlanta, GA 30303, USA.
Biochem Biophys Res Commun. 2004 Feb 6;314(2):622-30. doi: 10.1016/j.bbrc.2003.12.129.
We previously reported ATPase, RNA unwinding, and RNA-binding activities of recombinant p68 RNA helicase that was expressed in Escherichia coli. Huang et al. The recombinant protein bound both single-stranded (ss) and double-stranded (ds) RNAs. To further characterize the substrate RNA binding by p68 RNA helicase, we expressed and purified the recombinant N-terminal and C-terminal domains of the protein. RNA-binding property and protein phosphorylation of the recombinant domains of p68 were analyzed. Our data demonstrated that the C-terminal domain of p68 RNA helicase bound ssRNA. More interestingly, the C-terminal domain was a target of protein kinase C (PKC). Phosphorylation of the C-terminal domain of p68 abolished its RNA binding. Based on our observations, we propose that the C-terminal domain is an RNA substrate binding site for p68. The protein phosphorylation by PKC regulates the RNA binding of p68 RNA helicase, which consequently controls the enzymatic activities of the protein.
我们之前报道了在大肠杆菌中表达的重组p68 RNA解旋酶的ATP酶、RNA解旋和RNA结合活性。黄等人。该重组蛋白能结合单链（ss）和双链（ds）RNA。为了进一步表征p68 RNA解旋酶与底物RNA的结合，我们表达并纯化了该蛋白的重组N端和C端结构域。分析了p68重组结构域的RNA结合特性和蛋白磷酸化情况。我们的数据表明，p68 RNA解旋酶的C端结构域能结合单链RNA。更有趣的是，C端结构域是蛋白激酶C（PKC）的作用靶点。p68 C端结构域的磷酸化使其丧失RNA结合能力。基于我们的观察结果，我们提出C端结构域是p68的RNA底物结合位点。PKC介导的蛋白磷酸化调节p68 RNA解旋酶的RNA结合，进而控制该蛋白的酶活性。
