HslV-HslU: A novel ATP-dependent protease complex in Escherichia coli related to the eukaryotic proteasome
HslV-HslU: A novel ATP-dependent protease complex in Escherichia coli related to the eukaryotic proteasome
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DOI：
10.1073/pnas.93.12.5808
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发表时间：
1996-06-11
影响因子：
11.1
通讯作者：
Goldberg, AL
中科院分区：
文献类型：
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作者：
Rohrwild, M;Coux, O;Goldberg, AL
我们从大肠杆菌中分离到一种新的ATP依赖性蛋白酶，它是热休克位点hsIVU的产物，编码两种蛋白：HsIV，一种19-kDa的类似于蛋白酶体β亚基的蛋白，和HsIU，一种50-kDa的与ATP酶ClpX相关的蛋白。在ATP存在下，该蛋白酶能迅速水解荧光肽Z-Gly-Gly-Leu-AMC，而对某些胰凝乳蛋白酶底物的水解则非常缓慢。coli中组成型表达热休克蛋白，并且在从高拷贝质粒表达HsIV和BsIU的细胞中100倍。虽然HsIV和HsIU可以coimmunoprecipitated从两个菌株的细胞提取物与抗-HSIV抗体，这两个组件很容易通过各种类型的色谱分离。ATP刺激肽酶活性高达150倍，而其他核苷三磷酸，nonhydrolizable ATP类似物，ADP，或AMP没有影响。肽酶活性被阻断的抗HsIV抗体和几种类型的抑制剂的真核蛋白酶体（苏氨酸蛋白酶），但不是由其他类的蛋白酶的抑制剂，不像真核蛋白酶体，HsIVU蛋白酶缺乏胰蛋白酶样和肽基-谷氨酰-肽酶的活动。电子显微照片显示类似于20 S蛋白酶体或ClpAP蛋白酶的正面图像的环形颗粒。因此，HsIV和HsIU似乎形成复合物，其中HsIU的ATP水解对于肽水解bg（蛋白酶体样组分HsIV）是必需的。
We have isolated a new type of ATP-dependent protease from Escherichia coli, It is the product of the heat-shock Locus hsIVU that encodes two proteins: HsIV, a 19-kDa protein similar to proteasome beta subunits, and HsIU, a 50-kDa protein related to the ATPase ClpX. In the presence of ATP, the protease hydrolyzes rapidly the fluorogenic peptide Z-Gly-Gly-Leu-AMC and very slowly certain other chymotrypsin substrates, This activity increased 10-fold in E. coli expressing heat-shock proteins constitutively and 100-fold in cells expressing HsIV and BsIU from a high copy plasmid. Although HsIV and HsIU could be coimmunoprecipitated from cell extracts of both strains with an anti-HSIV antibody, these two components were readily separated by various types of chromatography. ATP stimulated peptidase activity up to 150-fold, whereas other nucleoside triphosphates, a nonhydrolzable ATP analog, ADP, or AMP had no effect. Peptidase activity was blocked by the anti-HsIV antibody and by several types of inhibitors of the eukaryotic proteasome (a threonine protease) but not by inhibitors of other classes of proteases, Unlike eukaryotic proteasomes, the HsIVU protease lacked tryptic-like and peptidyl-glutamyl-peptidase activities. Electron micrographs reveal ring-shaped particles similar to en face images of the 20S proteasome or the ClpAP protease. Thus, HsIV and HsIU appear to form a complex in which ATP hydrolysis by HsIU is essential for peptide hydrolysis bg,the proteasome-like component HsIV.