Nicoll D A, Quednau B D, Qui Z, Xia Y R, Lusis A J, Philipson K D
Department of Physiology, University of California, Los Angeles School of Medicine, Los Angeles, California 90095-1760, USA.
J Biol Chem. 1996 Oct 4;271(40):24914-21. doi: 10.1074/jbc.271.40.24914.
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.
NCX3是哺乳动物钠钙交换体中第三个被克隆的亚型。通过聚合酶链反应（PCR），利用源自NCX1和NCX2两个保守区域序列的简并引物，从大鼠脑cDNA中鉴定出NCX3。NCX3的PCR产物被用于从大鼠脑cDNA文库中分离出两个重叠克隆，总计4.8千碱基（kb）。对这两个重叠克隆进行测序，并在一个独特的Bsp106I限制性酶切位点处连接，形成一个全长cDNA克隆。NCX3 cDNA克隆有一个2.8 kb的开放阅读框，编码一个含927个氨基酸的蛋白质。在氨基酸水平上，NCX3与NCX1有73%的同一性，与NCX2有75%的同一性，预计其膜拓扑结构与NCX1和NCX2相同。在NCX3克隆上添加poly(A)+尾后，交换体活性可在非洲爪蟾卵母细胞中表达。NCX3也在哺乳动物BHK细胞系中表达。NCX3转录本大小为6 kb，高度局限于脑和骨骼肌。小鼠的连锁分析表明，NCX基因家族是分散的，因为NCX1、NCX2和NCX3基因分别定位于小鼠的17号、7号和12号染色体上。
