Expression of three members of the calcium-dependent protein kinase gene family in Arabidopsis thaliana
Expression of three members of the calcium-dependent protein kinase gene family in Arabidopsis thaliana
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DOI：
10.1007/bf00019557
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发表时间：
1996-03-01
影响因子：
5.1
通讯作者：
Chua, NH
中科院分区：
文献类型：
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作者：
Hong, Y;Takano, M;Chua, NH
钙依赖性蛋白激酶（CDPKs）属于一类独特的酶家族，其含有在氨基末端具有激酶结构域的单链和在羧基末端具有推定的钙结合EF手结构的单链多肽链。我们从拟南芥中克隆了三个编码CDPKs的cDNA序列，分别命名为atcdpk 6、ntcdpk 9和atcdpk 19。atcdpk 6、atcdpk 9和atcdpk 19的全长cDNA序列分别编码分子量为59343、55376和59947的蛋白质。在CDPK6和在CDPK9蛋白的重组是完全活性的激酶，其活性由Ca 2+诱导。生物化学研究表明，在激酶结构域和EF手结构之间的连接处存在自抑制结构域。在CDPK6的四个EF手的连续缺失表明，四个EF手的完整性是至关重要的钙反应。所有三个atcdpk基因在植物中普遍表达的RNA凝胶印迹实验证明。基因组序列的比较表明，三个cdpk基因的进化不同。使用针对CDPK6和CDPK9的抗体进行免疫组织化学实验，发现CDPKs以时间和发育调节的方式在特定细胞类型中表达。
Calcium-dependent protein kinases (CDPKs) belong to a unique family of enzymes containing a single polypeptide chain with a kinase domain at the amino terminus and a putative calcium-binding EF hands structure at the carboxyl terminus. From Arabidopsis thaliana, we have cloned three distinct cDNA sequences encoding CDPKs, which were designated as atcdpk6, ntcdpk9 and atcdpk19. The full-length cDNA sequences for atcdpk6, atcdpk9 and atcdpk19 encode proteins with a molecular weight of 59343, 55376 and 59947, respectively. Recombinant at CDPK6 and at CDPK9 proteins were fully active as kinases whose activities were induced by Ca2+. Biochemical studies suggested the presence of an autoinhibitory domain in the junction between the kinase domain and the EF hands structure. Serial deletion of the four EF hands of at CDPK6 demonstrated that the integrity of the four EF hands was crucial to the Ca2+ response. All the three atcdpk genes were ubiquitously expressed in the plant as demonstrated by RNA gel blot experiments. Comparison of the genomic sequences suggested that the three cdpk genes have evolved differently. Using antibodies against at CDPK6 and at CDPK9 for immunohistochemical experiments, CDPKs were found to be expressed in specific cell types in a temporally and developmentally regulated manner.