Nishida S, Fujita T, Kohno N, Atoda H, Morita T, Takeya H, Kido I, Paine M J, Kawabata S, Iwanaga S
Department of Biology, Faculty of Science, Kyushu University, Fukuoka, Japan.
Biochemistry. 1995 Feb 7;34(5):1771-8. doi: 10.1021/bi00005a034.
The complete amino acid sequence of ecarin is deduced from the nucleotide sequence of a cDNA clone isolated by screening a venomous gland cDNA library of Kenyan Echis carinatus. The cDNA sequence with 2379 base pairs encodes an open reading frame of 616 amino acids with a remarkable sequence homology to the putative precursor protein of trigramin from Trimeresurus gramineus venom (61% identity) and a large hemorrhagin, jararhagin, from the pit viper Bothrops jararaca venom (62% identity). Thus, ecarin, as well as jararhagin and trigramin, is translated as a precursor protein, which may be processed posttranslationally. The ecarin proprotein has a "cysteine switch" motif (-Pro-Lys-Met-Cys-Gly-Val-) similar to that involved in the activation of matrix metalloproteinase zymogens. The processed mature protein consists of 426 amino acid residues (residues 191-616), showing the strongest sequence similarity with that of Russell's viper venom factor X activator (RVV-X) heavy chain (64% identity). Like RVV-X heavy chain, ecarin contains metalloproteinase, disintegrin, and cysteine-rich domains. The metalloproteinase domain has a typical zinc-chelating sequence (-His-Glu-Xaa-Xaa-His-Xaa-Xaa-Gly-Xaa-Xaa-His-), as found in crayfish astacin. In the disintegrin domain of ecarin, the Arg-Gly-Asp sequence is replaced by Arg-Asp-Asp, as found in the disintegrin domains of RVV-X heavy chain (Arg-Asp-Glu) and a guinea pig sperm fusion protein, PH-30 beta (Thr-Asp-Glu). These findings show that while there are structural and evolutionary relationships among these proteins, each has a unique functional activity.
通过筛选肯尼亚锯鳞蝰毒腺cDNA文库分离得到的一个cDNA克隆的核苷酸序列，推导得出了伊卡林的完整氨基酸序列。该2379个碱基对的cDNA序列编码一个616个氨基酸的开放阅读框，与竹叶青蛇毒中的trigramin假定前体蛋白具有显著的序列同源性（同一性为61%），与蝮蛇Bothrops jararaca毒液中的一种大型出血毒素——矛头蝮毒素（同一性为62%）也有显著同源性。因此，伊卡林以及矛头蝮毒素和trigramin都是以前体蛋白形式翻译的，可能在翻译后进行加工。伊卡林前体蛋白具有一个“半胱氨酸开关”基序（-Pro-Lys-Met-Cys-Gly-Val-），类似于参与基质金属蛋白酶酶原激活的基序。加工后的成熟蛋白由426个氨基酸残基组成（第191 - 616位残基），与罗素蝰蛇毒因子X激活剂（RVV-X）重链的序列相似性最强（同一性为64%）。与RVV-X重链一样，伊卡林包含金属蛋白酶、去整合素和富含半胱氨酸的结构域。金属蛋白酶结构域具有典型的锌螯合序列（-His-Glu-Xaa-Xaa-His-Xaa-Xaa-Gly-Xaa-Xaa-His-），如在小龙虾的虾红素中发现的那样。在伊卡林的去整合素结构域中，Arg-Gly-Asp序列被Arg-Asp-Asp取代，如在RVV-X重链的去整合素结构域（Arg-Asp-Glu）和豚鼠精子融合蛋白PH-30β（Thr-Asp-Glu）中发现的那样。这些发现表明，虽然这些蛋白质之间存在结构和进化关系，但每种蛋白质都具有独特的功能活性。
