Heat Shock Factor 1 (HSF1) is the master regulator of the heat shock response, a universal survival mechanism throughout eukaryotic species used to buffer potentially lethal proteotoxic conditions. HSF1's function in vivo is regulated by several factors, including post translational modifications and elevated temperatures, whereupon it forms trimers to bind with heat shock elements in DNA. Unsurprisingly, HSF1 is also extremely sensitive to elevated temperatures in vitro, which poses specific technical challenges when producing HSF1 using a recombinant expression system. Although there are several useful publications which outline steps taken for HSF1 expression and purification, studies that describe specific strategies and detailed protocols to overcome HSF1 trimerisation and degradation are currently lacking. Herein, we have reported our detailed experimental protocol for the expression and purification of monomeric human HSF1 (HsHSF1) as a major species. We also propose a refined method of inducing HsHSF1 activation in vitro, that we consider more accurately mimics HsHSF1 activation in vivo and is therefore more physiologically relevant.

热休克因子1（HSF1）是热休克反应的主要调节剂，它是整个真核生物的普遍生存机制，用于缓冲可能致命的蛋白毒性疾病。HSF1在体内的功能受多种因素调节，包括翻译后修饰和温度升高，然后形成三聚体以与DNA中的热激元件结合。毫不奇怪，HSF1对体外高温也极为敏感，使用重组表达系统生产HSF1时，这会带来特定的技术挑战。尽管有一些有用的出版物概述了HSF1表达和纯化的步骤，但目前尚缺乏描述克服HSF1三聚和降解的具体策略和详细方案的研究。在这里，我们已经报告了作为主要物种的单体人HSF1（Hs HSF1）的表达和纯化的详细实验方案。我们还提出了一种体外诱导Hs HSF1激活的改进方法，我们认为它可以更准确地模拟体内Hs HSF1的激活，因此在生理上更相关。