Forrester Michael T, Thompson J Will, Foster Matthew W, Nogueira Leonardo, Moseley M Arthur, Stamler Jonathan S
Department of Biochemistry, Duke University Medical Center, Durham, North Carolina, USA.
Nat Biotechnol. 2009 Jun;27(6):557-9. doi: 10.1038/nbt.1545. Epub 2009 May 31.
We have modified the biotin switch assay for protein S-nitrosothiols (SNOs), using resin-assisted capture (SNO-RAC). Compared with existing methodologies, SNO-RAC requires fewer steps, detects high-mass S-nitrosylated proteins more efficiently, and facilitates identification and quantification of S-nitrosylated sites by mass spectrometry. When combined with iTRAQ labeling, SNO-RAC revealed that intracellular proteins may undergo rapid denitrosylation on a global scale. This methodology is readily adapted to analyzing diverse cysteine-based protein modifications, including S-acylation.
我们使用树脂辅助捕获法(SNO-RAC)对蛋白质S-亚硝基硫醇(SNOs)的生物素开关分析方法进行了改进。与现有方法相比,SNO-RAC所需步骤更少,能更有效地检测高质量的S-亚硝基化蛋白质,并且有助于通过质谱法鉴定和定量S-亚硝基化位点。当与iTRAQ标记相结合时,SNO-RAC显示细胞内蛋白质可能在全球范围内经历快速的去亚硝基化。该方法很容易适用于分析包括S-酰化在内的各种基于半胱氨酸的蛋白质修饰。