Characterizing the Covalent Targets of a Small Molecule Inhibitor of the Lysine Acetyltransferase P300
Characterizing the Covalent Targets of a Small Molecule Inhibitor of the Lysine Acetyltransferase P300
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DOI：
10.1021/acsmedchemlett.5b00385
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发表时间：
2016-02-01
影响因子：
4.2
通讯作者：
Meier, Jordan L.
中科院分区：
文献类型：
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作者：
Shrimp, Jonathan H.;Sorum, Alexander W.;Meier, Jordan L.
C646抑制赖氨酸乙酰转移酶（KATs）p300和CBP，是迄今为止鉴定出的最有效和最具选择性的小分子KAT抑制剂。为了深入了解这种表观遗传学探针的细胞活性，我们应用化学蛋白质组学来鉴定C646化学型的共价靶点。通过建模和合成衍生化，开发了一种可点击的类似物（C646 - 炔烃），它对p300的抑制作用与母体化合物相似，并能够富集结合的蛋白质。液相色谱 - 串联质谱（LC - MS/MS）鉴定出C646 - 炔烃的主要共价靶点是高丰度的含半胱氨酸蛋白质，后续研究发现C646在体外可抑制微管蛋白聚合。最后，我们提供的证据表明，C646的巯基反应性可能限制其在细胞中拮抗乙酰化的能力。这些发现应能更准确地解释将C646用作KAT活性化学探针的研究，并表明含亲电体的抑制剂一个未被充分认识的缺陷是，由于被大量蛋白质和代谢物的巯基库消耗，其细胞效力降低。
C646 inhibits the lysine acetyltransferases (KATs) p300 and CBP and represents the most potent and selective small molecule KAT inhibitor identified to date. To gain insights into the cellular activity of this epigenetic probe, we applied chemoproteomics to identify covalent targets of the C646 chemotype. Modeling and synthetic derivatization was used to develop a clickable analogue (C646-yne) that inhibits p300 similarly to the parent compound and enables enrichment of bound proteins. LC-MS/MS identified the major covalent targets of C646-yne as highly abundant cysteine-containing proteins, and follow-up studies found that C646 can inhibit tubulin polymerization in vitro. Finally, we provide evidence that thiol reactivity of C646 may limit its ability to antagonize acetylation in cells. These findings should enable a more precise interpretation of studies utilizing C646 as a chemical probe of KAT activity and suggest that an underappreciated liability of electrophile-containing inhibitors is a reduction in their cellular potency due to consumption by abundant protein and metabolite thiol sinks.