Functional Analysis of an ADAMTS10 Signal Peptide Mutation in Weill-Marchesani Syndrome Demonstrates a Long-Range Effect on Secretion of the Full-Length Enzyme
Functional Analysis of an ADAMTS10 Signal Peptide Mutation in Weill-Marchesani Syndrome Demonstrates a Long-Range Effect on Secretion of the Full-Length Enzyme
复制标题
DOI：
10.1002/humu.20797
复制
发表时间：
2008-12-01
期刊：
影响因子：
3.9
通讯作者：
Apte, Suneel S.
中科院分区：
文献类型：
--
作者：
Kutz, Wendy E.;Wang, Lauren W.;Apte, Suneel S.
我们报告了第一个错义ADAMTS 10突变（c.73G > A; p.Ala25Thr）的鉴定和功能分析，该突变导致隐性Weill-Marchesani综合征（WMS）。受错义突变影响的Ala 25残基位于相对于ADAMTS 10信号肽酶切割位点的-1位置。p.Ala25Thr取代的全长ADAMTS 10在HEK 293 F和Cos-1细胞中均显示出一致且显著减少的分泌。然而，缺乏辅助结构域和仅含有信号肽、前肽和催化结构域的C-末端截短的构建体（p.Ala25Thr Pro-Cat）在HEK 293 F细胞和Cos-1细胞中均有效分泌。纯化的p.Ala25Thr Pro-Cat和来自HEK 293 F细胞的p.Ala25Thr取代的全长ADAMTS 10的Edman降解证明了正确的信号肽加工。因此，p.Ala25Thr取代阻碍全长ADAMTS 10从细胞中分泌，但不阻碍Pro-Cat从细胞中分泌，但允许去除信号肽。我们推断，复杂的C-末端辅助结构域的折叠是ADAMTS 10生物合成的速率限制步骤，并且它（而不是Pro-Cat）对信号肽切割效率的细微变化敏感。这些观察结果代表了信号肽突变的前所未有的影响，并支持蛋白质生物合成过程中的初始共翻译加工事件对蛋白质折叠和分泌产生长期影响的模型。《Mutat》29（12），1425-1434，2008年。(C)2008 Wiley-Liss，Inc.
We report the identification and functional analysis of the first missense ADAMTS10 mutation (c.73G > A; p.Ala25Thr) causing recessive Weill-Marchesani syndrome (WMS). The Ala25 residue affected by the missense mutation is at the -1 position relative to the ADAMTS10 signal peptidase cleavage site. p.Ala25Thr substituted full-length ADAMTS10 showed consistent and significantly diminished secretion in both HEK293F and Cos-1 cells. However, a C-terminally truncated construct lacking the ancillary domain and containing only the signal peptide, the propeptide and the catalytic domain (p.Ala25Thr Pro-Cat) was efficiently secreted in both HEK293F cells and Cos-1 cells. Edman degradation of purified p.Ala25Thr Pro-Cat and p.Ala25Thr substituted full-length ADAMTS10 from HEK293F cells demonstrated correct signal peptide processing. Thus, the p.Ala25Thr substitution hinders secretion of full-length ADAMTS10, but not Pro-Cat from cells, yet permits signal peptide removal. We infer that folding of the complex C-terminal ancillary domain is the rate, limiting step in biosynthesis of ADAMTS10, and that it (but not Pro-Cat) is sensitive to subtle changes in efficiency of signal peptide cleavage. These observations represent an unprecedented effect of a signal peptide mutation and support a model in which the initial cotranslational processing events during protein biosynthesis can have long-range effects on protein folding and secretion. Hum Mutat 29(12), 1425-1434, 2008. (C) 2008 Wiley-Liss, Inc.