Systematic Mapping of Posttranslational Modifications in Human Estrogen Receptor-α with Emphasis on Novel Phosphorylation Sites
Systematic Mapping of Posttranslational Modifications in Human Estrogen Receptor-α with Emphasis on Novel Phosphorylation Sites
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DOI：
10.1074/mcp.m800282-mcp200
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发表时间：
2009-03-01
影响因子：
7
通讯作者：
Baldwin, Michael A.
中科院分区：
文献类型：
--
作者：
Atsriku, Christian;Britton, David J.;Baldwin, Michael A.
本文对人乳腺癌MCF-7细胞株雌激素受体的翻译后修饰进行了系统研究。用多种酶、质谱和串联质谱进行蛋白水解，获得了雌二醇处理的细胞培养物中全长66-kDa内源性蛋白的非常高的序列覆盖率。鉴定了9个磷酸化丝氨酸残基，其中3个以前未报道，没有一个以前由任何其他实验室通过质谱法观察到。在重组蛋白中鉴定了另外两个修饰的丝氨酸残基，一个先前报道但在内源性蛋白中未观察到，另一个先前未知。虽然主要重点放在确定新的磷酸化位点，N-末端损失的蛋氨酸伴随着氨基乙酰化和赖氨酸侧链乙酰化（或可能三甲基化）也被检测到。使用HPLC-ESI和MALDI连接到不同的质量分析仪得到更高的序列覆盖率，并确定了更多的网站比可以实现的任何一种方法单独。雌激素受体在乳腺癌的发生和发展中至关重要。一个以前未报道的磷酸化位点在这里被证明是强烈依赖于雌二醇，证实其对乳腺癌的潜在意义。更多地了解雌激素受体的翻译后修饰，特别是磷酸化，将增加我们对导致雌二醇诱导的这种蛋白质激活的过程的理解，并可能有助于开发用于管理雌激素依赖性乳腺癌的治疗策略。Molecular & Cellular Proteomics 8：467-480，2009.
A systematic study of posttranslational modifications of the estrogen receptor isolated from the MCF-7 human breast cancer cell line is reported. Proteolysis with multiple enzymes, mass spectrometry, and tandem mass spectrometry achieved very high sequence coverage for the full-length 66-kDa endogenous protein from estradiol-treated cell cultures. Nine phosphorylated serine residues were identified, three of which were previously unreported and none of which were previously observed by mass spectrometry by any other laboratory. Two additional modified serine residues were identified in recombinant protein, one previously reported but not observed here in endogenous protein and the other previously unknown. Although major emphasis was placed on identifying new phosphorylation sites, N-terminal loss of methionine accompanied by amino acetylation and a lysine side chain acetylation (or possibly trimethylation) were also detected. The use of both HPLC-ESI and MALDI interfaced to different mass analyzers gave higher sequence coverage and identified more sites than could be achieved by either method alone. The estrogen receptor is critical in the development and progression of breast cancer. One previously unreported phosphorylation site identified here was shown to be strongly dependent on estradiol, confirming its potential significance to breast cancer. Greater knowledge of this array of posttranslational modifications of estrogen receptor, particularly phosphorylation, will increase our understanding of the processes that lead to estradiol-induced activation of this protein and may aid the development of therapeutic strategies for management of hormone-dependent breast cancer. Molecular & Cellular Proteomics 8:467-480, 2009.