A disulfide bond is required for functional assembly of NCX1 from complementary fragments.
A disulfide bond is required for functional assembly of NCX1 from complementary fragments.
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从互补片段功能性组装 NCX1 需要二硫键。
DOI：
10.1006/bbrc.2001.5656
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发表时间：
2001
期刊：
影响因子：
--
通讯作者：
Philipson,KD
中科院分区：
文献类型：
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作者：
Qiu,Z;Chen,J;Nicoll,DA;Philipson,KD
心脏Na+-Ca 2+交换器由具有两个跨膜区段（TMS）簇的单个多肽组成，所述跨膜区段（TMS）簇由TMS 5和TMS 6之间的大细胞内环隔开（Nicoll等人（1999）J.Biol.Chem.274，910-917; Iwamoto等人（1999）FEBS Lett. 446，264-268）。可通过将交换器cDNA分成两个片段来表达“分裂”交换器，使得蛋白质的NH 2-和CO2 H-末端部分在HEK 293细胞中表达为单独的多肽。部分交换分子的表达没有导致可检测的交换活性。然而，共表达交换器的两个部分的细胞显示出完整的野生型交换器的活性的30%至50%。全长交换剂在残基14或20和792之间含有二硫键。我们通过诱变和表达研究研究了这种二硫键在分裂交换器中的作用。我们的研究结果表明，交换器的功能需要两个TMS簇和C（14或20）/C792二硫键是必不可少的活性交换器的表达从半分子。
The cardiac Na+–Ca2+exchanger consists of a single polypeptide with two transmembrane segment (TMS) clusters separated by a large intracellular loop between TMS5 and TMS6 (Nicoll et al. (1999) J. Biol. Chem. 274, 910–917; Iwamoto et al. (1999) FEBS Lett. 446, 264–268). A “split” exchanger can be expressed by dividing the exchanger cDNA into two fragments so that the NH2- and CO2H-terminal portions of the protein are expressed as separate polypeptides in HEK293 cells. Expression of partial exchanger molecules did not result in detectable exchanger activity. Cells coexpressing both portions of the exchanger, however, displayed between 30 and 50% of the activity of the intact wild-type exchanger. The full-length exchanger contains a disulfide bond between residues 14 or 20 and 792. We examined the role of this disulfide bond in the split exchanger by mutagenesis and expression studies. Our results indicate that the function of the exchanger requires both TMS clusters and that the C(14 or 20)/C792 disulfide bond is essential for expression of active exchangers from half molecules.