Imamura M, Sakurai T, Ogawa Y, Ishikawa T, Goto K, Masaki T
Institute of Basic Medical Sciences, University of Tsukuba, Ibaraki, Japan.
Eur J Biochem. 1994 Jul 15;223(2):395-401. doi: 10.1111/j.1432-1033.1994.tb19006.x.
We previously reported the purification and characterization of a novel non-muscle alpha-actinin from chicken lung [Imamura, M. & Masaki, T, (1992) J. Biol. Chem. 267, 25927-25933]. The Ca2+ sensitivity of the lung alpha-actinin for the interaction with polymerized actin (F-actin) was much lower than those of the other reported non-muscle alpha-actinins. Here, we isolated a cDNA clone encoding the novel alpha-actinin by screening a chicken lung lambda g11 cDNA library with antibody specific for the low-Ca(2+)-sensitive alpha-actinin. The deduced amino acid sequence of the lung alpha-actinin showed 76%, 82% and 83% identity to those of chicken skeletal muscle, smooth-muscle and fibroblast-type alpha-actinin, respectively. Marked difference in the structure between the lung-type and the other alpha-actinins was found in the extreme NH2-terminal and in the COOH-terminal half; in the third and fourth regions of four spectrin-like repeats, and in two Ca(2+)-binding EF-hand consensus regions. The NH2-terminal-side EF-hand contained a notable defect in one of the five oxygen-containing amino acid side chains involved in chelating Ca2+, suggesting that the lower Ca2+ sensitivity of the lung alpha-actinin is ascribable to this defect. Northern blot analysis showed that the expression pattern of lung-type alpha-actinin mRNA in various non-muscle tissues differed from that of the other known non-muscle-type (fibroblast-type) alpha-actinin. The present results clearly demonstrate the existence of two structurally and functionally different types of non-muscle alpha-actinin; high-Ca(2+)-sensitive-type (NM1) and low-Ca(2+)-sensitive-type (NM2) alpha-actinin.
我们之前报道过从鸡肺中纯化并鉴定出一种新型非肌肉α-辅肌动蛋白[今村，M. & 正木，T，(1992)《生物化学杂志》267, 25927 - 25933]。肺α-辅肌动蛋白与聚合肌动蛋白（F-肌动蛋白）相互作用的Ca²⁺敏感性远低于其他已报道的非肌肉α-辅肌动蛋白。在此，我们通过用对低Ca²⁺敏感性α-辅肌动蛋白特异的抗体筛选鸡肺λgt11 cDNA文库，分离出了编码该新型α-辅肌动蛋白的cDNA克隆。推导的肺α-辅肌动蛋白氨基酸序列与鸡骨骼肌、平滑肌和成纤维细胞型α-辅肌动蛋白的氨基酸序列分别具有76%、82%和83%的同一性。在肺型和其他α-辅肌动蛋白之间，在极端NH₂末端和COOH末端的一半区域；在四个血影蛋白样重复序列的第三和第四区域；以及在两个Ca²⁺结合EF手基序区域发现了结构上的显著差异。NH₂末端侧的EF手在参与螯合Ca²⁺的五个含氧氨基酸侧链之一中存在明显缺陷，这表明肺α-辅肌动蛋白较低的Ca²⁺敏感性归因于此缺陷。Northern印迹分析表明，肺型α-辅肌动蛋白mRNA在各种非肌肉组织中的表达模式与其他已知非肌肉型（成纤维细胞型）α-辅肌动蛋白的不同。目前的结果清楚地证明了存在两种结构和功能不同的非肌肉α-辅肌动蛋白类型；高Ca²⁺敏感型（NM1）和低Ca²⁺敏感型（NM2）α-辅肌动蛋白。
