美国加州大学Jason E. Gestwicki团队近期取得重要工作进展，他们研究提出了使用构象响应染料的蛋白质自适应差示扫描荧光法。相关研究成果2024年5月14日在线发表于《自然—生物技术》杂志上。

据介绍，差示扫描荧光法（DSF）是一种通过荧光染料选择性识别未折叠状态来报告蛋白质热稳定性的技术。然而，DSF的应用仍然受到蛋白质与现有DSF染料不相容性的限制。

研究人员通过开发蛋白质适应性DSF平台（paDSF）克服了这一障碍，该平台将染料库“Aurora”与简化的程序相结合，可根据需要识别蛋白质-染料对。paDSF成功应用于94%（70种蛋白质中的66种）的蛋白质，使之前的兼容性增加了两倍，并提供了66种功能和生化多样性蛋白质的分析，其中包括10种来自严重急性呼吸综合征冠状病毒2型的蛋白质。研究人员发现，paDSF可以用于监测以前无法访问的生物过程，证明了O-GlcNAc转移酶的结构域间异源性。

总之，Aurora染料的化学多样性和不同的选择性表明，paDSF功能可以很容易地扩展。paDSF是研究蛋白质稳定性、动力学和配体结合的通用工具。

附：英文原文

Title: Protein-adaptive differential scanning fluorimetry using conformationally responsive dyes

Author: Wu, Taiasean, Yu, Joshua C., Suresh, Arundhati, Gale-Day, Zachary J., Alteen, Matthew G., Woo, Amanda S., Millbern, Zoe, Johnson, Oleta T., Carroll, Emma C., Partch, Carrie L., Fourches, Denis, Vinueza, Nelson R., Vocadlo, David J., Gestwicki, Jason E.

Issue&Volume: 2024-05-14

Abstract: Differential scanning fluorimetry (DSF) is a technique that reports protein thermal stability via the selective recognition of unfolded states by fluorogenic dyes. However, DSF applications remain limited by protein incompatibilities with existing DSF dyes. Here we overcome this obstacle with the development of a protein-adaptive DSF platform (paDSF) that combines a dye library ‘Aurora’ with a streamlined procedure to identify protein–dye pairs on demand. paDSF was successfully applied to 94% (66 of 70) of proteins, tripling the previous compatibility and delivering assays for 66 functionally and biochemically diverse proteins, including 10 from severe acute respiratory syndrome coronavirus 2. We find that paDSF can be used to monitor biological processes that were previously inaccessible, demonstrated for the interdomain allostery of O-GlcNAc transferase. The chemical diversity and varied selectivities of Aurora dyes suggest that paDSF functionality may be readily extended. paDSF is a generalizable tool to interrogate protein stability, dynamics and ligand binding.

DOI: 10.1038/s41587-024-02158-7

Source: https://www.nature.com/articles/s41587-024-02158-7