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捕获蛋白质与蛋白质相互作用与酸性氨基酸反应性交叉链接器

Qing-Qing Liao^1,2, Xin Shu¹, Wei Sun¹

¹Zhejiang Provincial Key Laboratory for Cancer Molecular Cell Biology, Life Sciences Institute Cancer Center, Zhejiang University, Hangzhou, Zhejiang, 310058, China.

Small (Weinheim an der Bergstrasse, Germany)

|December 11, 2023

概括

新的交叉连接器针对蛋白质表面的酸性残留物,使得在中性pH下能够进行近距离增强的反应. 这种方法有效地识别了蛋白质-蛋白质相互作用,推进了结构生物学和蛋白质组学.

科学领域:

生物化学生物化学
蛋白质组学是指蛋白质组学.
结构生物学结构生物学

背景情况:

酸性残留物 (酸和胺酸) 在蛋白质表面上很常见,但针对它们的化学交叉链接具有挑战性.
针对酸性残留物的现有方法通常需要高度的试剂或表现出不良的结构兼容性.

研究的目的:

开发新型的异构功能交叉链接剂,可以与蛋白质表面的酸性残留物 (Asp和Glu) 反应.
建立一个与中性pH值和室温条件相容的近距离增强交叉连接策略.
扩大交联质谱法 (XL-MS) 的实用性,用于研究涉及酸性域的蛋白-蛋白相互作用.

主要方法:

扩展"种植和造"战略,以创建异性生物功能交叉链接器.
交叉连接剂与素侧链的序列反应,其次是酸和胺酸侧链.
在各种模型蛋白质和蛋白质复合体中证明交联效率.
应用交叉连接质谱法 (XL-MS) 来研究p53蛋白相互作用.

主要成果:

开发了异性生物功能交叉链接器,这些交叉链接器可以与Lys侧链快速反应,然后以近距离增强的方式与Asp/Glu侧链反应.
交叉连接反应在中性pH值和室温下有效地进行,无需合试剂.
从小单体到大复合体,在各种蛋白质类型中表现出强度.
使用XL-MS识别了p53C终端域的87个假定结合因子,包括SARNP,ZRAB2和WBP11,它们调节p53基因表达和拼接.

结论:

新型的碳酸盐反应交叉连接器为向酸性残留物提供了一种高效且结构兼容的方法.
这种方法显著提高了XL-MS在绘制蛋白质-蛋白质相互作用的能力,特别是涉及酸性域的蛋白质相互作用.
鉴定到的p53相互作用体突显了这种方法在发现与基因调节相关的功能性蛋白质关联方面的潜力.

关键词:

酸性氨基酸交叉链接的酸性氨基酸.化学交叉连接器化学交叉连接器交叉链接质谱仪的质谱仪.在 p53 里,p53 是一个 p53 的类型.

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