Top-down targeted proteomics for deep sequencing of tropomyosin isoforms.
Top-down targeted proteomics for deep sequencing of tropomyosin isoforms.
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DOI：
10.1021/pr301054n
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发表时间：
2013-01-04
影响因子：
4.4
通讯作者：
Ge Y
中科院分区：
文献类型：
--
作者：
Peng Y;Chen X;Zhang H;Xu Q;Hacker TA;Ge Y
原肌球蛋白（tromyosins, Tm）是一类普遍存在且高度保守的肌动蛋白结合蛋白，在多种生物过程中发挥重要作用。由多个Tm编码基因和外显子产生的Tm异构体以及翻译后修饰（PTMs）调节Tm的功能。因此，为了更好地了解Tm的功能作用，有必要充分表征Tm同工异构体。在此，我们开发了一种自上而下的基于高分辨率质谱（MS）的靶向蛋白质组学方法，用于全面表征Tm亚型。α-Tm是猪心肌的主要亚型。我们进一步对其序列进行了表征，并定位了其乙酰化、磷酸化和氨基酸多态性等PTMs。有趣的是，我们发现了一种“新颖的”Tm异构体，它与任何现有的猪Tm序列都不匹配。自上而下质谱法对该亚型进行了深度测序，发现该亚型与小鼠β-Tm序列完全匹配，表明该“新型”亚型是猪β-Tm，在猪和小鼠之间100%保守。综上所述，我们证明了自上而下的靶向蛋白质组学提供了一种强大的工具，可以从遗传变异中对Tm亚型进行深度测序，并对PTM位点进行完整的定位。
Tropomyosins (Tm) constitute a family of ubiquitous and highly conserved actin-binding proteins, playing essential roles in a variety of biological processes. Tm isoforms produced by multiple Tm encoding genes and alternatively expressed exons along with post-translational modifications (PTMs) regulate Tm function. Therefore, to gain a better understanding of the functional role of Tm, it is essential to fully characterize Tm isoforms. Herein, we developed a top-down high-resolution mass spectrometry (MS) based targeted proteomics method for comprehensive characterization of Tm isoforms. α–Tm was identified to be the predominant isoform in swine cardiac muscle. We further characterized its sequence and localized the PTMs such as acetylation and phosphorylation as well as amino acid polymorphisms. Interestingly, we discovered a “novel” Tm isoform that does not match with any of the currently available swine Tm sequences. A deep sequencing of this isoform by top-down MS revealed an exact match with mouse β–Tm sequence, suggesting that this “novel” isoform is swine β–Tm which is 100% conserved between swine and mouse. Taken together, we demonstrated that top-down targeted proteomics provides a powerful tool for deep sequencing of Tm isoforms from genetic variations together with complete mapping of the PTM sites.
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影响因子：
29.4
作者：
Lin, JLC;Geng, X;Lin, JJC
通讯作者：
Lin, JJC
影响因子：
8.8
作者：
Franzen, B;Linder, S;Auer, G
通讯作者：
Auer, G
DOI：
10.1073/pnas.75.8.3588
发表时间：
1978-01-01
影响因子：
11.1
作者：
MAK, A;SMILLIE, LB;BARANY, M
通讯作者：
BARANY, M
DOI：
10.1016/j.bbrc.2004.06.084
发表时间：
2004-08-06
影响因子：
3.1
作者：
Denz, CR;Narshi, A;Dube, DK
通讯作者：
Dube, DK
影响因子：
6.2
作者：
Hughes, JAI;Cooke-Yarborough, CM;Weinberger, RP
通讯作者：
Weinberger, RP