Functional dissection of the PE domain responsible for translocation of PE_PGRS33 across the mycobacterial cell wall.
Functional dissection of the PE domain responsible for translocation of PE_PGRS33 across the mycobacterial cell wall.
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DOI：
10.1371/journal.pone.0027713
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发表时间：
2011
期刊：
影响因子：
3.7
通讯作者：
Manganelli R
中科院分区：
文献类型：
--
作者：
Cascioferro A;Daleke MH;Ventura M;Donà V;Delogu G;Palù G;Bitter W;Manganelli R
PE 是一种特殊的输出分枝杆菌蛋白，在致病性分枝杆菌物种中含量过高。它们的特征是具有约 110 个氨基酸的 N 端结构域（PE 结构域），已被证明负责它们的输出和定位。在本文中，我们描述了 PE_PGRS33 (PERv1818c) 的 PE 结构域，它是特征最明确的 PE 蛋白之一。我们构建了几种突变蛋白，其中 PE 结构域的部分被删除或发生确定的突变。这些蛋白质在不同的分枝杆菌物种中表达，并对其定位进行了表征。我们证实 PE 结构域对于 PE_PGRS33 表面定位至关重要，并证明缺少前 30 个氨基酸的 PE 结构域会失去其功能。然而，所有 PE 蛋白 N 端结构域内两个极其保守的区域中的单个氨基酸取代对 PE_PGRS33 的稳定性有一定影响，但对其定位没有影响。使用海分枝杆菌，我们可以证明 VII 型分泌系统 ESX-5 对于 PE_PGRS33 输出至关重要。此外，在海分枝杆菌中，但在牛分枝杆菌 BCG 和结核分枝杆菌中，PE_PGRS33 的 PE 结构域在缺乏 PGRS 结构域的情况下表达时会被加工并分泌到培养基中。最后，使用将 PERv1818c 结构域的不同部分融合到绿色荧光蛋白 N 末端的嵌合蛋白，我们可以假设 PE 结构域的前 30 个氨基酸包含允许蛋白质易位的序列。
PE are peculiar exported mycobacterial proteins over-represented in pathogenic mycobacterial species. They are characterized by an N-terminal domain of about 110 amino acids (PE domain) which has been demonstrated to be responsible for their export and localization. In this paper, we characterize the PE domain of PE_PGRS33 (PERv1818c), one of the best characterized PE proteins. We constructed several mutated proteins in which portions of the PE domain were deleted or subjected to defined mutations. These proteins were expressed in different mycobacterial species and their localization was characterized. We confirmed that the PE domain is essential for PE_PGRS33 surface localization, and demonstrated that a PE domain lacking its first 30 amino acids loses its function. However, single amino acid substitutions in two regions extremely well conserved within the N-terminal domain of all PE proteins had some effect on the stability of PE_PGRS33, but not on its localization. Using Mycobacterium marinum we could show that the type VII secretion system ESX-5 is essential for PE_PGRS33 export. Moreover, in M. marinum, but not in Mycobacterium bovis BCG and in Mycobacterium tuberculosis, the PE domain of PE_PGRS33 is processed and secreted into the culture medium when expressed in the absence of the PGRS domain. Finally, using chimeric proteins in which different portions of the PERv1818c domain were fused to the N-terminus of the green fluorescent protein, we could hypothesize that the first 30 amino acids of the PE domain contain a sequence that allows protein translocation.
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影响因子：
6.7
作者：
Sani M;Houben EN;Geurtsen J;Pierson J;de Punder K;van Zon M;Wever B;Piersma SR;Jiménez CR;Daffé M;Appelmelk BJ;Bitter W;van der Wel N;Peters PJ
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Peters PJ
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4.8
作者：
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通讯作者：
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