Tsumoto Hiroki, Akimoto Yoshihiro, Endo Tamao, Miura Yuri
Research Team for Mechanism of Aging, Tokyo Metropolitan Institute of Gerontology, 35-2 Sakae-cho, Itabashi-ku, Tokyo 173-0015, Japan.
Department of Anatomy, Kyorin University School of Medicine, 6-20-2 Shinkawa, Mitaka-shi, Tokyo 181-8611, Japan.
Bioorg Med Chem Lett. 2017 Nov 15;27(22):5022-5026. doi: 10.1016/j.bmcl.2017.10.005. Epub 2017 Oct 4.
Protein O-GlcNAcylation regulates various biological processes, and is associated with several diseases. Therefore, the development of quantitative proteomics is important for understanding the mechanisms of O-GlcNAc-related diseases. We previously reported selective enrichment of O-GlcNAcylated peptides, which provided high-selectivity and effective release by a novel thiol-alkyne and thiol-disulfide exchange. Here, we describe a new approach using initial isobaric tag labeling for relative quantification followed by enrichment and β-elimination/Michael addition with dithiothreitol for identification of both proteins and modification sites. The approach was validated using model proteins and peptides. This novel strategy could be used for quantitative O-GlcNAcome of biological samples.
蛋白质O-连接的N-乙酰葡糖胺化修饰调控多种生物学过程，并与多种疾病相关。因此，定量蛋白质组学的发展对于理解O-连接的N-乙酰葡糖胺相关疾病的机制至关重要。我们之前报道了O-连接的N-乙酰葡糖胺化肽段的选择性富集，其通过新型硫醇-炔烃和硫醇-二硫键交换提供了高选择性和有效释放。在此，我们描述了一种新方法，该方法首先使用等压标签标记进行相对定量，然后进行富集以及与二硫苏糖醇的β-消除/迈克尔加成反应，以鉴定蛋白质及其修饰位点。该方法使用模型蛋白质和肽段进行了验证。这种新策略可用于生物样品的O-连接的N-乙酰葡糖胺组定量分析。
