HeLa细胞在紫外线辐射条件下的磷酸化质谱数据集
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为了更好地了解细胞对DNA损伤的反应，我们用高灵敏度的质谱法对HeLa细胞的磷酸化进行了比较性的蛋白质组学研究。共确定了2100个蛋白质的4367个磷酸化位点，其中许多位点以前未曾报道过。 收获的 ""重 ""和 ""轻 ""标记的细胞分别用2×NETN缓冲液（200mM NaCl, 100mM Tris-Cl, 2mM EDTA, 1.0 % NP-40, pH 7.2）在冰上裂解30分钟。在4℃下20,000×g离心10分钟后，保存上清液。所得的MS/MS数据用MaxQuant和集成的Andromeda搜索引擎（v.1.4.1.2）处理。串联质谱数据在SwissProt_human数据库（20,274个序列）和反向诱饵数据库进行了搜索。裂解酶使用的是Trypsin/P，允许最多2个缺失的裂解，每条肽有4个修饰和5个电荷。前体离子的质量误差为20ppm，碎片离子的质量误差为0.02Da。半胱氨酸上的氨基甲基化被指定为固定修饰，蛋氨酸上的氧化，丝氨酸上的磷酸化被指定为可变修饰。蛋白质、肽和修饰位点的错误发现率（FDR）阈值为1%。MaxQuant中的所有其他参数保持默认值。
To better understand the cellular response to DNA damage, we conducted a comparative proteomic study of phosphorylation in HeLa cells using high-sensitivity mass spectrometry. A total of 4367 phosphorylation sites across 2100 proteins were identified, many of which have not been previously reported. Harvested "heavy" and "light" labeled cells were lysed on ice for 30 minutes using 2×NETN buffer (200 mM NaCl, 100 mM Tris-Cl, 2 mM EDTA, 1.0% NP-40, pH 7.2). After centrifugation at 20,000×g for 10 minutes at 4°C, the supernatant was collected and stored. The resulting MS/MS data were processed using MaxQuant and the integrated Andromeda search engine (v.1.4.1.2). Tandem mass spectrometry data were searched against the SwissProt_human database (containing 20,274 sequences) and a reversed decoy database. Trypsin/P was employed as the proteolytic enzyme, with a maximum of 2 missed cleavages permitted, up to 4 modifications and 5 charge states allowed per peptide. The mass tolerance for precursor ions was set to 20 ppm, while that for fragment ions was 0.02 Da. Carbamidomethylation on cysteine was designated as a fixed modification, whereas oxidation on methionine and phosphorylation on serine were designated as variable modifications. The false discovery rate (FDR) threshold for proteins, peptides, and modification sites was set to 1%. All other parameters within MaxQuant were retained as their default settings.
