Hung C H, Huang H R, Huang C J, Huang F L, Chang G D
Graduate Institute of Biochemical Sciences, National Taiwan University, Taipei 10098, Taiwan.
J Biol Chem. 1997 May 23;272(21):13772-8. doi: 10.1074/jbc.272.21.13772.
We have purified a secreted proteinase of 23 kDa from carp head kidney by sequential column chromatography on a Reactive Blue 72-agarose dye affinity column and an FPLC Mono-P column. The secretion of this proteinase from carp head kidney can be stimulated by high concentrations of potassium. Since the carp proteinase is present mainly in the head kidney, kidney, and spleen (all of which are lymphohematopoietic organs), it is named nephrosin. The carp nephrosin is most sensitive to metal chelators, but not to inhibitors specific for other classes of proteinases. A cDNA clone has been isolated from a carp head kidney cDNA library by immunoscreening with a polyclonal antiserum raised against purified nephrosin. The cloned cDNA is 1086 base pairs in length and has an open reading frame encoding a protein of 273 amino acids, including a 19-amino acid signal peptide and 56-amino acid propeptide. The deduced amino acid sequence shows moderate levels of identity to medaka HCE1 (52.5%), medaka LCE (50.7%), crayfish astacin (33.2%), murine meprin-alpha (34%), and murine meprin-beta (33.5%), all members of the astacin family of zinc endopeptidases. Nephrosin is the first member of the astacin family found in lymphohematopoietic tissues.
我们通过在活性蓝72 - 琼脂糖染料亲和柱和FPLC单P柱上进行连续柱层析，从鲤鱼头肾中纯化出了一种23 kDa的分泌型蛋白酶。高浓度钾可刺激这种蛋白酶从鲤鱼头肾中分泌。由于鲤鱼蛋白酶主要存在于头肾、肾脏和脾脏（均为淋巴造血器官）中，故将其命名为肾素。鲤鱼肾素对金属螯合剂最为敏感，但对其他类蛋白酶的特异性抑制剂不敏感。通过用针对纯化肾素产生的多克隆抗血清进行免疫筛选，从鲤鱼头肾cDNA文库中分离出了一个cDNA克隆。克隆的cDNA长度为1086个碱基对，有一个开放阅读框，编码一个273个氨基酸的蛋白质，包括一个19个氨基酸的信号肽和一个56个氨基酸的前肽。推导的氨基酸序列与青鳉HCE1（52.5%）、青鳉LCE（50.7%）、小龙虾虾红素（33.2%）、小鼠meprin - alpha（34%）和小鼠meprin - beta（33.5%）有中等程度的同源性，它们都是锌内肽酶虾红素家族的成员。肾素是在淋巴造血组织中发现的虾红素家族的首个成员。
