Quantification of mitochondrial S-nitrosylation by CysTMT⁶ switch assay.
Quantification of mitochondrial S-nitrosylation by CysTMT⁶ switch assay.
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DOI：
10.1007/978-1-62703-386-2_14
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发表时间：
2013
影响因子：
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通讯作者：
Christopher I Murray;H. S. Chung;H. Uhrigshardt;J. V. Van Eyk
中科院分区：
文献类型：
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作者：
Christopher I Murray;H. S. Chung;H. Uhrigshardt;J. V. Van Eyk
S-亚硝基化（SNO）是一种重要的心肌线粒体氧化翻译后修饰。几种线粒体蛋白的SNO修饰与缺血/再灌注损伤后心脏预处理和改善细胞存活相关。由于其不稳定的性质，SNO修饰是具有挑战性的研究使用传统的生物化学技术，特别是，个别修饰的半胱氨酸残基的鉴定。在这里，我们描述的cysTMT 6开关检测，经典的生物素开关协议的变化的细节。cysTMT 6试剂通过结合修饰的半胱氨酸残基的明确鉴定和质谱分析最多6个样品的相对定量，提供了一种简单而强大的SNO检测方法。
S-nitrosylation (SNO) is an important oxidative posttranslational modification in the regulation of cardiac mitochondria. SNO modification of several mitochondrial proteins has been associated with cardiac preconditioning and improved cell survival following ischemia/reperfusion injury. Due to their labile nature, SNO modifications are challenging to study using traditional biochemical techniques; particularly, the identification of individual modified cysteine residues. Here, we describe the details of the cysTMT6switch assay, a variation of the classic biotin switch protocol. The cysTMT6reagent provides a simplified and powerful approach to SNO detection by combining unambiguous identification of the modified cysteine residue and relative quantification of up to six samples by mass spectrometry analysis.