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Molecular mechanism ofreguLaled inframembrane proteolysis (RIP)

调节性膜内蛋白水解（RIP）的分子机制

基本信息

批准号：
17370068
负责人：
AKIYAMA Yoshinori
金额：
$ 10.22万
依托单位：
Kyoto University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2007
项目状态：
已结题

项目摘要

Regulated intramembrane proteolysis (RIP) plays key roles in the regulation of various cellular processes. During RIP, a target membrane protein is believed to receive limited cleavage within the transmembrane segment. E. coli has two RIP proteases, RseP and GlpG. We studied these proteins to understand their physiological roles and the mechanism of RIP. We found that RseP cleaved RseA, a negative regulator of transcription factor σ_E, within its transmembrane region and that RseP was thereby essential for the activation of the σ_E pathway of ESR. We showed that the active site of RseP is sequestered form the surrounding lipidic and aqueous environments and exists in a folded protein domain. We also characterized the proteolytic activity of RseP. Although the physiological function of GlpG remains unknown, we constructed a model substrate and demonstrated that GlpG possesses proteolytic activity against model membrane proteins. We showed that although the GlpG active site exists within a hydrophilic cavity open to the periplasm, which is consistent with the recently reported crystal structures of GlpG, it cleaves a substrate at a region located outside the membrane. We identified two sequence elements in a model substrate required for cleavage by GlpG and suggested a model of the protease action of GlpG. We also analyzed the structure and function of the L1 periplasmic loop of GlpG, and suggested that it protrudes from the GlpG core region and plays a regulatory role for the GlpG activity.

受调节的膜内蛋白水解（RIP）在各种细胞过程的调节中起着关键作用。在RIP过程中，靶膜蛋白被认为在跨膜区段内接受有限的切割。E.大肠杆菌有两种RIP蛋白酶，RseP和GlpG。我们研究了这些蛋白质，以了解它们的生理作用和RIP的机制。我们发现RseP在其跨膜区切割转录因子σ_E的负调节因子RseA，从而RseP是激活ESR的σ_E途径所必需的。我们发现RseP的活性位点与周围的水环境隔离，并存在于折叠的蛋白质结构域中。我们还表征了RseP的蛋白水解活性。虽然GlpG的生理功能仍然未知，我们构建了一个模型底物，并证明GlpG具有对模型膜蛋白的蛋白水解活性。我们发现，虽然GlpG活性位点存在于一个亲水性的空腔开放的周质，这是一致的，最近报道的晶体结构的GlpG，它裂解基板位于膜外的区域。我们确定了两个序列元素中的模型底物所需的GlpG裂解，并提出了一个模型的GlpG的蛋白酶作用。我们还分析了GlpG的L1周质环的结构和功能，认为它突出于GlpG的核心区域，对GlpG的活性起调节作用。