Chemical proteomic profiling of protein N-homocysteinylation with a thioester probe†,Chemical Science

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Chemical proteomic profiling of protein N-homocysteinylation with a thioester probe†
Chemical Science ( IF 7.6 ) Pub Date : 2018-02-16 00:00:00 , DOI: 10.1039/c8sc00221e
Nan Chen _{1,

2,

3,

4,

5} , Jinmin Liu _{1,

2,

3,

4,

5} , Zeyu Qiao _{1,

2,

3,

4,

5} , Yuan Liu _{1,

2,

3,

4,

5} , Yue Yang _{5,

6,

7,

8,

9} , Changtao Jiang _{5,

6,

7,

8,

9} , Xian Wang _{5,

6,

7,

8,

9} , Chu Wang _{1,

2,

3,

4,

5}

Affiliation

Hyperhomocysteinemia (HHcy) refers to a medical condition of abnormally high level of homocysteine (Hcy) in blood (>15 μmol L⁻¹) and has been clinically implicated with cardiovascular diseases and neurodegenerative disorders. Excessive Hcy can be converted to a reactive thioester intermediate, Hcy thiolactone (HTL), which selectively reacts with protein lysine residues (“N-homocysteinylation”) and this non-enzymatic modification largely contributes to manifestations of HHcy. However, the proteome-wide detection of protein N-homocysteinylation remains a challenge to date. In this work, we report a chemoselective reaction to label and enrich N-homocysteinylation from complex proteome samples as inspired by native chemical ligation for protein synthesis. Alkynyl thioester probes are synthesized and the reaction is validated with small molecule and purified protein models successfully. We performed quantitative chemical proteomics to identify more than 800 N-homocysteinylated proteins as well as 304 N-homocysteinylated sites directly from HTL-treated HeLa cells. The chemical proteomics strategies will facilitate functional study of protein N-homocysteinylations in the HHcy-implicated diseases.

中文翻译：

用硫酯探针对蛋白质N-高半胱氨酸化的化学蛋白质组学分析†

高同型半胱氨酸血症（HHcy）是指血液中高半胱氨酸（Hcy）异常高水平（> 15μmolL ^-1）的医学状况，在临床上与心血管疾病和神经退行性疾病有关。过量的Hcy可转化为反应性硫酯中间体Hcy硫代内酯（HTL），可与蛋白质赖氨酸残基选择性反应（“ N-同型半胱氨酸化”），这种非酶修饰很大程度上有助于HHcy的表现。然而，迄今为止，蛋白质组全蛋白N-同型半胱氨酸化的检测仍然是一个挑战。在这项工作中，我们报告了标记和富集氮的化学选择性反应蛋白质组合成的自然化学连接的启发，从复杂的蛋白质组样本中提取了半胱氨酸。合成了炔基硫酯探针，并成功地用小分子和纯化的蛋白质模型验证了反应。我们进行了定量化学蛋白质组学研究，直接从HTL处理的HeLa细胞中鉴定出800多个N-同型半胱氨酸化的蛋白质以及304个N-同型半胱氨酸化的位点。化学蛋白质组学策略将有助于在涉及HHcy的疾病中蛋白质N-同型半胱氨酸化的功能研究。

更新日期：2018-02-16

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