Analysis of tubulin alpha-1A/1B C-terminal tail post-translational poly-glutamylation reveals novel modification sites.
Analysis of tubulin alpha-1A/1B C-terminal tail post-translational poly-glutamylation reveals novel modification sites.
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DOI：
10.1021/pr2011044
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发表时间：
2012-03-02
影响因子：
4.4
通讯作者：
Byers, Stephen W.
中科院分区：
文献类型：
--
作者：
Sahab, Ziad J.;Kirilyuk, Alexander;Zhang, Lihua;Khamis, Zahraa I.;Pompach, Petr;Sung, YouMe;Byers, Stephen W.
关键词：
微管蛋白-α1A/1B C-末端尾部在其序列的最后11个氨基酸中有7个谷氨酸残基，它们是潜在的谷氨酸化位点。另一个主要的修饰是C-末端酪氨酸的裂解导致微管蛋白-α1A/1B的降解形式。除其他外，这些修饰在脑细胞和微管中产生了高度异质性的微管蛋白样本，在指导细胞内运输、微管动力学和有丝分裂事件方面发挥了重要作用，并可能因细胞和疾病状态而异，如癌症和神经退行性疾病。之前使用初级质谱仪(MS)离子和部分埃德曼测序进行了鉴定，仅在E445残基上发现了微管蛋白-α1A/1B谷氨酰化。我们描述了2DE分离、胰酶和蛋白酶K凝胶内消化以及小鼠脑和牛微管的纳米α-ESI-QqTOF-MS/MS分析微管蛋白-ESI-1B的谷氨酰化和酪氨酸化。通过比较内源性和合成肽的碎裂模式和保留时间，鉴定了酪氨酸化、去酪氨酸化和Δ2-微管蛋白-α1A/1B CTT。严格的验收标准被用来鉴定新的谷氨酰化位点。除了先前在E445上发现的位点外，在E441和E443上还检测到小鼠和牛微管蛋白-α1A/1B CTTS上的谷氨酰化，吉祥物期望值低于0.01。谷氨酸的O-甲基化也被观察到。
Tubulin-α1A/1B C-terminal tail (CTT) has 7 glutamic acid residues among the last 11 amino acids of its sequence that are potential sites for glutamylation. Cleavage of C-terminal tyrosine resulting in the detyrosinated form of tubulin-α1A/1B is another major modification. These modifications among others bring about highly heterogeneous tubulin samples in brain cells and microtubules, play a major role in directing intracellular trafficking, microtubule dynamics, and mitotic events, and can vary depending on the cell and disease state, such as cancer and neurodegenerative disorders. Identified previously using primary mass spectrometry (MS) ions and partial Edman sequencing, tubulin-α1A/1B glutamylation was found exclusively on the E445 residue. We here describe the analysis of tubulin-α1A/1B glutamylation and detyrosination after 2DE separation, trypsin and proteinase K in-gel digestion, and nanoUPLC-ESI-QqTOF-MS/MS of mouse brain and bovine microtubules. Tyrosinated, detyrosinated, and Δ2- tubulin-α1A/1B CTTs were identified based on a comparison of fragmentation patterns and retention times between endogenous and synthetic peptides. Stringent acceptance criteria were adapted for the identification of novel glutamylation sites. In addition to the previously identified site at E445, glutamylation on mouse and bovine tubulin-α1A/1B CTTs was identified on E441 and E443 with MASCOT Expect values below 0.01. O-methylation of glutamates was also observed.
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影响因子：
64.5
作者：
Chen H;Vermulst M;Wang YE;Chomyn A;Prolla TA;McCaffery JM;Chan DC
通讯作者：
Chan DC
影响因子：
7.4
作者：
Jung, Sung Yun;Li, Yehua;Qin, Jun
通讯作者：
Qin, Jun
影响因子：
1.8
作者：
Harrison, AG
通讯作者：
Harrison, AG
影响因子：
--
作者：
Khamis, Zahraa I.;Sahab, Ziad J.;Sang, Qing-Xiang Amy
通讯作者：
Sang, Qing-Xiang Amy
影响因子：
56.9
作者：
EDDE, B;ROSSIER, J;DENOULET, P
通讯作者：
DENOULET, P