Conserved Pro-Glu (PE) and Pro-Pro-Glu (PPE) Protein Domains Target LipY Lipases of Pathogenic Mycobacteria to the Cell Surface via the ESX-5 Pathway
Conserved Pro-Glu (PE) and Pro-Pro-Glu (PPE) Protein Domains Target LipY Lipases of Pathogenic Mycobacteria to the Cell Surface via the ESX-5 Pathway
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DOI：
10.1074/jbc.m110.204966
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发表时间：
2011-05-27
影响因子：
4.8
通讯作者：
Bitter, Wilbert
中科院分区：
文献类型：
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作者：
Daleke, Maria H.;Cascioferro, Alessandro;Bitter, Wilbert
VII型分泌系统ESX-5是致病性分枝杆菌中PE和PPE蛋白输出的主要途径。这些分枝杆菌特异性蛋白质家族的特征在于保守的100和180个氨基酸的N-末端结构域，其包含脯氨酸-谷氨酸（PE）和脯氨酸-脯氨酸-谷氨酸（PPE）基序，它们以其命名。在这里，我们研究了分泌的三酰甘油脂肪酶LipY，其中在快速生长的分枝杆菌包含一个信号序列，但在缓慢生长的物种似乎已经取代了信号肽与PE或PPE域。选择LipY同源物在野生型海洋分枝杆菌及其相应的ESX-5突变体中表达，并通过免疫印迹和电子显微镜研究蛋白质的定位。本研究表明，结核分枝杆菌PE-LipY（LipY（tub））和M. marinum PPE-LipY（LipY（mar））均以ESX-5依赖性方式分泌到细菌表面。转运后，PE/PPE结构域通过蛋白水解裂解除去。相反，具有信号序列的金黄分枝杆菌LipY不被转运到细胞表面。此外，我们表明，LipY（浴缸）和LipY（马尔）需要各自的PE和PPE域的ESX-5依赖性分泌。PE结构域在ESX-5分泌中的作用在全细胞脂肪酶测定中得到证实，其中表达全长LipY（tub）但不表达缺乏其PE结构域的LipY（tub）的野生型细菌显示水解细胞外脂质。总之，PE和PPE结构域都含有ESX-5系统分泌LipY所需的信号，并且这些结构域在易位时被蛋白水解除去。
The type VII secretion system ESX-5 is a major pathway for export of PE and PPE proteins in pathogenic mycobacteria. These mycobacteria-specific protein families are characterized by conserved N-terminal domains of 100 and 180 amino acids, which contain the proline-glutamic acid (PE) and proline-proline-glutamic acid (PPE) motifs after which they are named. Here we investigated secretion of the triacylglycerol lipase LipY, which in fast-growing mycobacteria contains a signal sequence, but in slow-growing species appears to have replaced the signal peptide with a PE or PPE domain. Selected LipY homologues were expressed in wild-type Mycobacterium marinum and its corresponding ESX-5 mutant, and localization of the proteins was investigated by immunoblotting and electron microscopy. Our study shows that Mycobacterium tuberculosis PE-LipY (LipY(tub)) and M. marinum PPE-LipY (LipY(mar)) are both secreted to the bacterial surface in an ESX-5-dependent fashion. After transport, the PE/PPE domains are removed by proteolytic cleavage. In contrast, Mycobacterium gilvum LipY, which has a signal sequence, is not transported to the cell surface. Furthermore, we show that LipY(tub) and LipY(mar) require their respective PE and PPE domains for ESX-5-dependent secretion. The role of the PE domain in ESX-5 secretion was confirmed in a whole cell lipase assay, in which wild-type bacteria expressing full-length LipY(tub), but not LipY(tub) lacking its PE domain, were shown to hydrolyze extracellular lipids. In conclusion, both PE and PPE domains contain a signal required for secretion of LipY by the ESX-5 system, and these domains are proteolytically removed upon translocation.