A plasminogen-like protein selectively degrades stearoyl-CoA desaturase in liver microsomes
A plasminogen-like protein selectively degrades stearoyl-CoA desaturase in liver microsomes
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DOI：
10.1074/jbc.m306240200
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发表时间：
2003-10-31
影响因子：
4.8
通讯作者：
Ozols, J
中科院分区：
文献类型：
--
作者：
Heinemann, FS;Korza, G;Ozols, J
硬脂酰辅酶A去饱和酶（stearoyl-CoA desaturase，SCD）是内质网膜上的一种蛋白质，在37 ℃孵育时能被选择性降解。我们以前报道了纯化的90 kDa的微粒体蛋白与SCD蛋白酶活性和其特征在于抑制剂的敏感性的蛋白酶。在这里，我们表明，90-kDa的蛋白质是一种微粒体形式的纤溶酶原（PG）和纯化的SCD蛋白酶含有一系列的纤溶酶样衍生物。90 kDa的蛋白质通过其胰蛋白酶肽的质谱鉴定为Pg。纯化的SCD蛋白酶与Pg抗体反应，免疫印迹证明富集Pg的SCD蛋白酶建立的纯化程序。通过酶谱分析的微粒体表明，在70 kDa的蛋白水解活性对应的流动性的Pg在非还原聚丙烯酰胺凝胶中的一个单一的带。当微粒体在酶谱分析之前在37 ℃下孵育时，在30 kDa处产生了强烈的蛋白水解活性条带。纯化的SCD蛋白酶显示范围从70至30 kDa的蛋白水解带谱。纯化的蛋白酶和微粒体对SCD的降解被来自药用水蛭水蛭的纤溶酶抑制剂bdellin抑制。为了探索Pg在体内SCD降解中的作用，我们研究了从Pg缺陷（Pg-/-）小鼠分离的微粒体中SCD的表达和降解。与野生型同窝对照小鼠的微粒体相比，Pg-/-小鼠的肝微粒体具有显著更高的SCD水平。Pg-/-小鼠微粒体中SCD的降解显著减少，而对照小鼠的肝微粒体显示出与大鼠肝微粒体中观察到的相似的快速SCD降解。这些发现表明SCD被与Pg相关的蛋白酶降解，并表明纤溶酶兼职作为细胞内蛋白酶。
Stearoyl-CoA desaturase (SCD) is an integral membrane protein of the endoplasmic reticulum that is rapidly and selectively degraded when isolated liver microsomes are incubated at 37degreesC. We previously reported the purification of a 90-kDa microsomal protein with SCD protease activity and characterized the inhibitor sensitivity of the protease. Here we show that the 90-kDa protein is a microsomal form of plasminogen (Pg) and that the purified SCD protease contains a spectrum of plasmin-like derivatives. The 90-kDa protein was identified as Pg by mass spectrometry of its tryptic peptides. The purified SCD protease reacted with Pg antibody, and immunoblotting demonstrated enrichment of Pg by the purification procedure established for the SCD protease. Analysis of microsomes by zymography demonstrated a single band of proteolytic activity at 70-kDa corresponding to the mobility of Pg in nonreduced polyacrylamide gels. When microsomes were incubated at 37degreesC prior to zymography, an intense band of proteolytic activity developed at 30-kDa. The purified SCD protease displayed a spectrum of proteolytic bands ranging from 70 to 30 kDa. Degradation of SCD by the purified protease and by microsomes was inhibited by bdellin, a plasmin inhibitor from the medicinal leech Hirudo medicinalis. To explore the role of Pg in the degradation of SCD in vivo, we examined SCD expression and degradation in microsomes isolated from Pg-deficient (Pg-/-) mice. Compared with microsomes from wild-type littermate control mice, liver microsomes from Pg-/- mice had significantly higher levels of SCD. Degradation of SCD in microsomes from Pg-/- mice was markedly diminished, whereas liver microsomes from control mice showed rapid SCD degradation similar to that observed in rat liver microsomes. These findings indicate that SCD is degraded by a protease related to Pg and suggest that plasmin moonlights as an intracellular protease.