Cloning of a third mammalian Na+-Ca2+ exchanger, NCX3
Cloning of a third mammalian Na+-Ca2+ exchanger, NCX3
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DOI：
10.1074/jbc.271.40.24914
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发表时间：
1996-10-04
影响因子：
4.8
通讯作者：
Philipson, KD
中科院分区：
文献类型：
--
作者：
Nicoll, DA;Quednau, BD;Philipson, KD
NCX3是克隆的第三个哺乳动物Na+-Ca~(2+)交换异构体。利用来自NCX1和NCX2两个保守区序列的简并引物，通过聚合酶链式反应(PCR)从大鼠脑组织中鉴定出NCX3。用NCX3扩增产物从大鼠脑cDNA文库中分离出两个大小为4.8kb的重叠克隆。对重叠克隆进行测序，连接到Bsp106I酶切位点，形成全长cDNA克隆。NCX3克隆的开放阅读框为2.8kb，编码927个氨基酸的蛋白质。在氨基酸水平上，NCX3与NCX1有73%的同源性，与NCX2有75%的同源性，并预测与NCX1和NCX2具有相同的膜拓扑结构。在NCX3克隆中加入Poly(A)(+)尾巴后，交换器活性可以在非洲爪哇卵母细胞中表达。NCX3在哺乳动物BHK细胞中也有表达。NCX3转录本的大小为6kb，高度限制在大脑和骨骼肌中。在小鼠中的连锁分析表明，NCX基因家族是分散的，因为NCX1、NCX2和NCX3基因分别定位在小鼠的第17、7和12号染色体上。
NCX3 is the third isoform of a mammalian Na+-Ca2+ exchanger to be cloned. NCX3 was identified from rat brain cDNA by polymerase chain reaction (PCR) using degenerate primers derived from the sequences of two conserved regions of NCX1 and NCX2. The NCX3 PCR product was used to isolate two overlapping clones totalling 4.8 kilobases (kb) from a rat brain cDNA library. The overlapping clones were sequenced and joined at a unique Bsp106I restriction enzyme site to form a full-length cDNA clone. The NCX3 cDNA clone has an open reading frame of 2.8 kb encoding a protein of 927 amino acids. At the amino acid level, NCX3 shares 73% identity with NCX1 and 75% identity with NCX2 and is predicted to share the same membrane topology as NCX1 and NCX2. Following addition of a poly(A)(+) tail to the NCX3 clone, exchanger activity could be expressed in Xenopus oocytes. NCX3 was also expressed in the mammalian BHK cell line. NCX3 transcripts are 6 kb in size and are highly restricted to brain and skeletal muscle. Linkage analysis in the mouse indicated that the NCX family of genes is dispersed, since the NCX1, NCX2, and NCX3 genes mapped to mouse chromosomes 17, 7, and 12, respectively.