The plasma membrane H+-ATPase from maize roots is phosphorylated in the C-terminal domain by a calcium-dependent protein kinase
The plasma membrane H+-ATPase from maize roots is phosphorylated in the C-terminal domain by a calcium-dependent protein kinase
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DOI：
10.1034/j.1399-3054.1998.1040405.x
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发表时间：
1998-12-01
影响因子：
6.4
通讯作者：
Aducci, P
中科院分区：
文献类型：
--
作者：
Camoni, L;Fullone, MR;Aducci, P
一种与玉米根质膜相关的蛋白激酶活性进行了部分纯化和表征。生化特性，如钙依赖性，抑制钙调素拮抗剂，钙调素刺激的情况下，表明该酶属于钙依赖性蛋白激酶（CDPK）家族。通过凝胶内磷酸化测定法测定活性多肽的分子量：55和51 kDa的两条带被标记。同样的蛋白质也被抗大豆CDPK的单克隆抗体免疫修饰。从体外试验的结果表明，玉米H+-ATP酶是一个合适的底物，这种蛋白激酶和磷酸化位点位于C-末端结构域的酶。这一结果得到证实，通过使用作为底物在磷酸化试验中的分离的C-末端结构域的H+-ATP酶在大肠杆菌中表达的谷胱甘肽-转移酶融合蛋白。
A protein kinase activity associated with maize root plasma membranes was partially purified and characterized. Biochemical properties, such as calcium dependence, inhibition by calmodulin antagonists, and absence of calmodulin stimulation; indicated that the enzyme belongs to the calcium-dependent protein kinase (CDPK) family. By means of an in-gel phosphorylation assay the molecular mass of active polypeptides was determined: two bands of 55 and 51 kDa became labelled. The same proteins were also immunodecorated by monoclonal antibodies against soybean CDPK. Results from in vitro assays demonstrated that maize H+-ATPase was a suitable substrate for this protein kinase and that the phosphorylation site was located at the C-terminal domain of the enzyme. This result was confirmed by using as substrate in phosphorylation assays the isolated C-terminal domain of the H+- ATPase expressed in Escherichia coli as a glutathione-transferase fusion protein.