Deletion and site-directed mutagenesis of EF-hand domain of phospholipase C-delta 1: effects on its activity.
Deletion and site-directed mutagenesis of EF-hand domain of phospholipase C-delta 1: effects on its activity.
复制标题
磷脂酶 C-delta 1 EF-hand 结构域的缺失和定点突变：对其活性的影响。
DOI：
--
复制
发表时间：
1995
期刊：
影响因子：
--
通讯作者：
Y. Nozawa
中科院分区：
文献类型：
--
作者：
S. Nakashima;Y. Banno;Takehito Watanabe;Yoshikatsu Nakamura;T. Mizutani;Hideki Sakai;Yutong Zhao;Y. Sugimoto;Y. Nozawa
为了阐明磷脂酶C-δ 1（PLC-δ 1）中一个推定的EF-手形基序（144-172）的作用，对该酶进行了缺失和点突变，并在CHO细胞和E. coliAD 202菌株。缺失141-236或173-236的氨基酸残基导致PLC活性的消除。然而，通过删除EF-手形基序（144-172）使PLC活性降低至15-20%仍然是Ca（2+）依赖性的。此外，突变体，其中保守的Asp 153，Asp 157，Glu 164或所有这些酸性氨基酸EF-手基序被替换为丙氨酸残基，显示几乎相同的PLC活性和Ca（2+）-依赖性的野生型。这些结果表明，该区域包含EF-手基序可能不发挥作用，在调控钙（2+）-敏感性的PLC-δ 1，但它的活性是重要的。
In order to elucidate a role of a putative EF-hand motif (144-172) in phospholipase C-delta 1 (PLC-delta 1), deletion and point mutation of the enzyme were performed and the mutated cDNAs were expressed in CHO cells and E. coli AD202 strain. Deletion of amino acid residues of 141-236 or 173-236 resulted in abolition of PLC activity. However, the decreased PLC activity to 15-20% by deletion of the EF-hand motif (144-172) was still Ca(2+)-dependent. Furthermore, mutants, in which conserved Asp153, Asp157, Glu164 or all these acidic amino acids in the EF-hand motif were replaced with alanine residues, showed nearly the same PLC activity and Ca(2+)-dependency as those of wild-type. These results suggest that the region containing the EF-hand motif may not play a role in regulation of Ca(2+)-sensitivity of PLC-delta 1, but is important for its activity.