Proteome-wide Mapping of Endogenous SUMOylation Sites in Mouse Testis.
Proteome-wide Mapping of Endogenous SUMOylation Sites in Mouse Testis.
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DOI：
10.1074/mcp.m116.062125
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发表时间：
2017-05
期刊：
影响因子：
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通讯作者：
He F
中科院分区：
文献类型：
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作者：
Cai L;Tu J;Song L;Gao Z;Li K;Wang Y;Liu Y;Zhong F;Ge R;Qin J;Ding C;He F
SUMO化是一种可逆的翻译后修饰，参与各种重要的生物学过程。迄今为止，存在有限的方法用于内源性野生型SUMO修饰的肽富集和SUMO化位点鉴定。在这项研究中，我们产生了一个高亲和力的SUMO 1抗体，以促进从胰蛋白酶/Lys-C蛋白酶消化的内源性SUMO 1修饰肽的富集。二次Glu-C蛋白酶消化后，我们确定了53个高置信SUMO 1修饰位点，从小鼠睾丸使用高分辨率质谱。生物信息学分析表明，SUMO 1修饰的蛋白质在转录调控和DNA修复中富集。Nab 1被证实是一个真正的SUMO化蛋白，Lys 479被鉴定为主要的SUMO化位点。Nab 1的SUMO化增强了其与HDAC 2的相互作用，并保持了其对EGR 1转录活性的抑制作用。因此，我们提供了一种新的方法来研究内源性SUMO化位点的组织样品。
SUMOylation is a reversible post-translational modification involved in various critical biological processes. To date, there is limited approach for endogenous wild-type SUMO-modified peptides enrichment and SUMOylation sites identification. In this study, we generated a high-affinity SUMO1 antibody to facilitate the enrichment of endogenous SUMO1-modified peptides from Trypsin/Lys-C protease digestion. Following secondary Glu-C protease digestion, we identified 53 high-confidence SUMO1-modified sites from mouse testis by using high-resolution mass spectrometry. Bioinformatics analyses showed that SUMO1-modified proteins were enriched in transcription regulation and DNA repair. Nab1 was validated to be an authentic SUMOylated protein and Lys479 was identified to be the major SUMOylation site. The SUMOylation of Nab1 enhanced its interaction with HDAC2 and maintained its inhibitory effect on EGR1 transcriptional activity. Therefore, we provided a novel approach to investigating endogenous SUMOylation sites in tissue samples.