Department of Pharmaceutical Technology, University of Turin, Turin, Italy.
UK Dementia Research Institute at the Maurice Wohl Institute of King's College London, London, UK.
Methods Mol Biol. 2021;2353:79-95. doi: 10.1007/978-1-0716-1605-5_5.
Iron-sulfur (Fe-S) clusters are key cofactors for proteins involved in essential cellular processes such as DNA replication and repair, ribosome biogenesis, tRNA thio-modification, and co-enzyme synthesis. Fe-S clusters can assemble spontaneously from inorganic compounds, but their biogenesis requires dedicated machineries to circumvent the toxic nature of iron and sulfur. To address how these machines work, different laboratories have applied various biochemical and biophysical approaches, both in vivo and in vitro. Fe-S cluster enzymatic and chemical formation in vitro is the most efficient way to follow Fe-S cluster biogenesis in a controlled environment and investigate each component of the machinery at the molecular level. In this review, we detail and discuss an efficient protocol for an in vitro Fe-S cluster enzymatic and chemical formation, which we successfully developed to study Fe-S cluster formation. We underline the applications of this approach to the study of an essential biological system.
铁硫 (Fe-S) 簇是参与重要细胞过程的蛋白质的关键辅因子，如 DNA 复制和修复、核糖体生物发生、tRNA 硫修饰和辅酶合成。Fe-S 簇可以从无机化合物中自发组装，但它们的生物发生需要专门的机器来规避铁和硫的毒性。为了解决这些机器如何工作的问题，不同的实验室已经在体内和体外应用了各种生化和生物物理方法。体外酶促和化学形成 Fe-S 簇是在控制环境中跟踪 Fe-S 簇生物发生并在分子水平上研究机器每个组件的最有效方法。在这篇综述中，我们详细讨论了一种有效的体外 Fe-S 簇酶促和化学形成方案，我们成功地开发了该方案来研究 Fe-S 簇的形成。我们强调了这种方法在研究重要生物系统中的应用。
