Identification of critical amino acid residues in the regulatory N-terminal domain of PMEL.
Identification of critical amino acid residues in the regulatory N-terminal domain of PMEL.
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DOI：
10.1038/s41598-021-87259-y
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发表时间：
2021-04-08
影响因子：
4.6
通讯作者：
Leonhardt RM
中科院分区：
文献类型：
--
作者：
Mitchell SM;Graham M;Liu X;Leonhardt RM
色素细胞特异性蛋白PMEL在黑素体中形成一个功能性淀粉样蛋白基质，色素黑色素沉积其上。淀粉样蛋白核心由一个短的蛋白水解片段组成，我们将其称为核心 - 淀粉样片段（CAF），可能还有该蛋白质的其他部分，比如PKD结构域。一个同样源自PMEL蛋白水解的高度O - 糖基化重复（RPT）结构域与淀粉样蛋白结合，并且对于形成聚集体的片状形态是必需的。调节性的N末端被排除在聚集体之外，然而它必须与CAF顺式连接以驱动淀粉样蛋白的形成。然后，该结构域可能在一个新的CAF亚基掺入新生淀粉样蛋白之前、期间或之后立即被切割掉。我们之前已经确定了一个21个氨基酸长的区域，它介导N末端对CAF的调节活性。然而，相应区段的许多突变导致错误折叠和/或阻断PMEL从内质网输出，使得它们的表型难以解释。在这里，我们采用一种以单氨基酸分辨率靶向该基序的饱和诱变方法。我们的结果证实了PMEL N末端区域的关键性质，并确定了几个对PMEL淀粉样蛋白生成至关重要的残基。
The pigment cell-specific protein PMEL forms a functional amyloid matrix in melanosomes onto which the pigment melanin is deposited. The amyloid core consists of a short proteolytic fragment, which we have termed the core-amyloid fragment (CAF) and perhaps additional parts of the protein, such as the PKD domain. A highly O-glycosylated repeat (RPT) domain also derived from PMEL proteolysis associates with the amyloid and is necessary to establish the sheet-like morphology of the assemblies. Excluded from the aggregate is the regulatory N-terminus, which nevertheless must be linked in cis to the CAF in order to drive amyloid formation. The domain is then likely cleaved away immediately before, during, or immediately after the incorporation of a new CAF subunit into the nascent amyloid. We had previously identified a 21 amino acid long region, which mediates the regulatory activity of the N-terminus towards the CAF. However, many mutations in the respective segment caused misfolding and/or blocked PMEL export from the endoplasmic reticulum, leaving their phenotype hard to interpret. Here, we employ a saturating mutagenesis approach targeting the motif at single amino acid resolution. Our results confirm the critical nature of the PMEL N-terminal region and identify several residues essential for PMEL amyloidogenesis.
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DOI：
10.1073/pnas.0904532106
发表时间：
2010-02-02
影响因子：
11.1
作者：
Friedrich, Ralf P.;Tepper, Katharina;Faendrich, Marcus
通讯作者：
Faendrich, Marcus
影响因子：
2.9
作者：
Brunberg E;Andersson L;Cothran G;Sandberg K;Mikko S;Lindgren G
通讯作者：
Lindgren G
DOI：
10.1083/jcb.200806081
发表时间：
2008-10-20
期刊：
The Journal of cell biology
影响因子：
--
作者：
Fritz R;Stiasny K;Heinz FX
通讯作者：
Heinz FX
影响因子：
9.8
作者：
Fowler DM;Koulov AV;Alory-Jost C;Marks MS;Balch WE;Kelly JW
通讯作者：
Kelly JW
影响因子：
4.4
作者：
Ehses, S;Leonhardt, RM;Knittler, MR
通讯作者：
Knittler, MR