一种具有皮科莫尔亲缘关系的O-GlcNAc转移酶多抑制剂的鉴定
Forrest A Hammel1, N Connor Payne2,3, Victoria M Marando1
1Department of Microbiology, Harvard Medical School, Boston, Massachusetts 02115, United States.
Journal of the American Chemical Society
|September 14, 2024
概括
O-GlcNAc转移酶 (OGT) 与宿主细胞因子1 (HCF-1) 结合在一起. 这种相互作用能够强烈抑制OGT活性,从而为细胞研究提供一种新的可逆性抑制剂.
科学领域:
- 生物化学
- 分子生物学
- 酵素学
背景情况:
- O-GlcNAc转移酶 (OGT) 是通过蛋白质糖化调节细胞过程的关键酶.
- 主细胞因子1 (HCF-1) 是已知的OGT基质和相互作用因子,但其与OGT的结合亲和力仍未定量.
- 了解OGT-HCF-1的相互作用是解读OGT的监管机制的关键.
研究的目的:
- 量化OGT及其主要结合伙伴HCF-1之间的结合亲和力.
- 阐明OGT-HCF-1相互作用的分子基础.
- 开发一种基于HCF-1结合动机的新型OGT抑制剂.
主要方法:
- 时间解析的Förster共振能量转移 (TR-FRET) 试验以测量OGT-连接体结合动力学.
- OGT的四重复 (TPR) 域的位点定向突变,以确定关键的结合残留物.
- 在体外和细胞测试以评估HCF-1衍生的OGT抑制.
- 开发一种基因编码,可诱导的OGT抑制剂.
主要成果:
- 一个HCF-1衍生的多 (HCF3R) 对OGT表现出皮克分子结合亲和力 (K_D ≤85 pM).
- 在OGT的TPR域内保存的青素对于高亲和度的HCF-1结合至关重要,突变会使亲和度降低5个数量级以上.
- 在体外和基于细胞的测试中,HCF3R强烈抑制了OGT活性.
- 一种新的,基因编码的,可诱导和可逆的OGT抑制剂已成功开发.
结论:
- OGT-HCF-1相互作用的特点是异常高的亲和力,由特定的分子接触驱动.
- 这种高亲和度的相互作用可以被利用来产生强大的OGT抑制剂.
- 开发的可诱导抑制剂为以可调和可逆的方式研究OGT功能提供了有价值的工具.
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