Aminopeptidase-A. I. CDNA cloning and expression and localization in rat tissues.
Aminopeptidase-A. I. CDNA cloning and expression and localization in rat tissues.
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氨肽酶-A。
DOI：
10.1152/ajpregu.2000.278.2.r413
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发表时间：
2000
期刊：
影响因子：
--
通讯作者：
Healy,DP
中科院分区：
文献类型：
--
作者：
Troyanovskaya,M;Jayaraman,G;Song,L;Healy,DP
氨肽酶-A（阿帕）是一种选择性水解寡肽氨基末端酸性残基的胞外酶，包括具有生物活性的[Asp 1]ANG II和[Asp 1]CCK-8。我们试图通过cDNA克隆和表达来表征大鼠阿帕，并通过原位杂交和免疫组织化学来确定其组织分布。从大鼠肾脏cDNA文库中分离的重叠cDNA克隆的序列分析表明，全长cDNA编码一个945个氨基酸的蛋白质，预测分子量为108 kDa的大小证实了在体外翻译的全长cDNA构建体。全长cDNA构建体在哺乳动物细胞中的瞬时转染产生了大小为140 kDa的蛋白质，该大小与来自大鼠组织的阿帕的免疫印迹一致，并且与阿帕被称为糖基化蛋白一致。组织阿帕活性和mRNA表达在肾脏和回肠中最高。原位杂交和免疫组化定位的阿帕表明，除了肾脏和回肠，阿帕分别定位于近端小管和肠上皮细胞的管腔刷状缘，阿帕与毛细血管或窦状隙的衬里。已知作为ANG II生理靶点的区域，包括肾小球、肾小球囊和垂体前叶，具有高水平的阿帕。定位模式表明阿帕可能有助于多种功能，即，在肽清除中的一般作用，以及在循环或局部产生的ANG II或CCK-8的代谢中的可能更具体的作用。
Aminopeptidase-A (APA) is an ectoenzyme that selectively hydrolyzes acidic residues from the amino terminus of oligopeptides, including biologically active [Asp1]ANG II and [Asp1]CCK-8. We sought to characterize rat APA by cDNA cloning and expression and to determine its tissue distribution by in situ hybridization and immunohistochemistry. Sequence analysis of overlapping cDNA clones isolated from rat kidney cDNA libraries indicated that the full-length cDNA encoded a 945-amino acid protein with a predicted molecular mass of 108 kDa; the size was confirmed by in vitro translation of a full-length cDNA construct. Transient transfection of the full-length cDNA construct in mammalian cells yielded a protein ∼140 kDa in size, a size that agrees with the immunoblots of APA from rat tissue and is consistent with APA being known as a glycosylated protein. Tissue APA activity and mRNA expression were highest in the kidney and ileum. Localization of APA by in situ hybridization and immunohistochemistry indicated that, with the exception of the kidney and ileum, where APA was localized to the luminal brush border of proximal tubules and enterocytes, respectively, APA was associated with either capillaries or the lining of sinusoids. Areas known to be physiological targets for ANG II, including glomeruli, the zona glomerulosa, and anterior pituitary, had high levels of APA. The localization pattern suggests that APA may subserve multiple functions, i.e., a generalized role in peptide scavenging and perhaps a more specific role in metabolism of circulating or locally produced ANG II or CCK-8.