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Isolation and characterizaion of renin processing enzyme from renin granule

肾素颗粒中肾素加工酶的分离和表征

基本信息

批准号：
04670567
负责人：
IKEDA Masaharu
金额：
$ 1.28万
依托单位：
University of Occupational & Environmental Health
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1992
资助国家：
日本
起止时间：
1992 至 1993
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-04670567/
关键词：
Renin Prorenin Processing Renin Granule Subtilisin プレセシング

项目摘要

Processing mechanism of prorenin to renin is still controversial. It is postulated that renin is synthesized and processed in the rough endoplasmic reticulum and the Golgi complex and stored in the renin granules and secreted by exocytosis. Recently several processing enzymes which process prohormones or protein precursors were found and identified as intrinsic processing enzymes.The purpose of this study was to clarify the nature of the intrinsic renin processing enzyme in the subcellular fraction of the kidney cortex. Generally prohormone convertases or protein precursor processing enzymes were reported to a common catalytic domain homologous to that of the subtilisin serine protease family.Hence in the first step of the present study, we investigated whether subtilisin per se can process prorenin to renin. When radiolabelled[^<35>S]-human recombinant prorenin was incubated with subtilisin BNP^-, human recombinant prorenin which showed a molecular weight of 43,000 dalton was cleaved … More to renin, a molecular weight of 38,000 dalton. In addition, when semipurified human amniotic prorenin(or inactive renin) was incubated with subtilisin, we confirmed the activation of prorenin by subtilisin by detection of active renin which was quantitated by a specific immunoradiometric assay system. These results indicate that subtilisin per se can process and activate prorenin to active form.In the next study, we investigated whether renin granule fraction of the kidney can process recombinant human prorenin to renin by cleaving prosegment of the prorenin. Human prorenin (molecular weight 43,000) was cleaved to renin (molecular weight 38,000) by renin granule fraction. This result suggests that renin granule fraction contained prorenin processing enzyme. In this study, we tried to isolate the prorenin processing enzyme from renin granule fraction, but we could noto isolate or identified prorenin processing enzyme in the renin granule because of loss of activity probably due to its lability during isolation. Less

原肾素加工成肾素的机制仍有争议。据推测，肾素在粗面内质网和高尔基复合体中合成和加工，储存在肾素颗粒中并通过胞吐分泌。近年来发现了几种加工激素原或蛋白质前体的加工酶，并鉴定为内源性加工酶，本研究旨在阐明肾皮质亚细胞组分中内源性肾素加工酶的性质。一般来说，激素原转化酶或蛋白前体加工酶被报道与枯草杆菌蛋白酶丝氨酸蛋白酶家族具有相同的催化结构域，因此在本研究的第一步，我们研究了枯草杆菌蛋白酶本身是否可以将原肾素加工成肾素。当放射性标记的[^<35>S]-人重组前肾素与枯草杆菌蛋白酶BNP^-孵育时，显示分子量为43，000道尔顿的人重组前肾素被裂解 ...更多信息转化为分子量为38,000道尔顿的肾素。此外，当半纯化的人羊膜原肾素（或失活的肾素）与枯草杆菌蛋白酶孵育时，我们通过检测由特定的免疫放射测定系统定量的活性肾素来证实枯草杆菌蛋白酶对原肾素的激活。这些结果表明，枯草杆菌蛋白酶本身可以处理和激活原肾素活性形式。在接下来的研究中，我们研究了是否可以处理重组人原肾素的原肾素的原片段切割的肾脏的肾素颗粒部分。人原肾素（分子量43，000）被肾素颗粒部分裂解成肾素（分子量38，000）。该结果表明，肾素颗粒级分含有原肾素加工酶。在本研究中，我们试图从肾素颗粒组分中分离出前肾素加工酶，但我们诺托分离或鉴定出肾素颗粒中的前肾素加工酶，因为可能是由于其在分离过程中的不稳定性而失去活性。少