Covalent Modification of Bromodomain Proteins by Peptides Containing a DNA Damage-Induced, Histone Post-Translational Modification.
Covalent Modification of Bromodomain Proteins by Peptides Containing a DNA Damage-Induced, Histone Post-Translational Modification.
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通过含有DNA损伤引起的翻译后修饰的肽对溴结构域蛋白的共价修饰。
DOI：
10.1002/cbic.202200373
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发表时间：
2022-11-18
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亲电性5-亚甲基-2-吡咯酮修饰（KMP）在核小体核心颗粒中的组蛋白的赖氨酸残基上与通常形成的DNA损伤（C4-AP）反应时产生。非酶促KMP修饰也在用抗肿瘤剂博来霉素处理的HeLa细胞的组蛋白中产生，博来霉素氧化DNA并形成C4-AP。这种非酶共价组蛋白修饰具有与N-乙酰赖氨酸（KAc）修饰相同的电荷，但更亲电。在这项研究中，我们表明，含有KMP的组蛋白肽被识别，并共价修饰溴结构域蛋白，是KAc读者。在与不同序列的含KMP的肽孵育后观察到共价溴结构域修饰的不同选择性偏好。3个共价修饰的溴结构域蛋白的MS/MS分析证实Cys加合是选择性的。修饰的Cys并不总是接近KAc结合位点，表明含KMP的肽与溴结构域蛋白的相互作用不同于前者。蛋白质加合产率作为溴结构域pH（蛋白质电荷为零（pI））或半胱氨酸溶剂可及表面积的函数的分析也与蛋白质和亲电子肽之间的非混杂相互作用一致。这些数据表明，KMP的细胞内形成可以通过修饰调节基因表达的蛋白质来影响细胞功能和活力。KMP修饰的组蛋白是由DNA损伤C4-AP产生的非酶翻译后修饰。化学合成的含KMP的组蛋白肽选择性地修饰含半胱氨酸的布罗莫结构域蛋白（N-乙酰赖氨酸的阅读器）。核蛋白，如溴结构域，通过KMP的共价修饰可以影响它们在细胞中的功能。
An electrophilic 5-methylene-2-pyrrolone modification (KMP) is produced at lysine residues of histone proteins in nucleosome core particles upon reaction with a commonly formed DNA lesion (C4-AP). The nonenzymatic KMP modification is also generated in the histones of HeLa cells treated with the antitumor agent, bleomycin that oxidizes DNA and forms C4-AP. This nonenzymatic covalent histone modification has the same charge as the N-acetyllysine (KAc) modification but is more electrophilic. In this study we show that KMP-containing histone peptides are recognized by, and covalently modify bromodomain proteins that are KAc readers. Distinct selectivity preferences for covalent bromodomain modification are observed following incubation with KMP-containing peptides of different sequence. MS/MS analysis of 3 covalently modified bromodomain proteins confirmed that Cys adduction was selective. The modified Cys was not always proximal to the KAc binding site, indicating that KMP-containing peptide interaction with bromodomain protein is distinct from the former. Analysis of protein adduction yields as a function of bromodomain pH at which the protein charge is zero (pI) or cysteine solvent accessible surface area are also consistent with non-promiscuous interaction between the proteins and electrophilic peptides. These data suggest that intracellular formation of KMP could affect cellular function and viability by modifying proteins that regulate genetic expression. KMP-modified histones are a nonenzymatic, post-translational modification produced from the DNA lesion C4-AP. Chemically synthesized KMP-containing histone peptides selectively modify cysteine-containing bromodomain proteins, the readers of N-acetyllysine. Covalent modification of nuclear proteins, such as bromodomains, by KMP could affect their function in cells.
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