Ferroportin (Fpn) is an essential mammalian iron transporter that is negatively regulated by the hormone hepcidin. Our current molecular understanding of Fpn‐mediated iron efflux and regulation is limited due to a lack of biochemical, biophysical and high‐resolution structural studies. A critical step towards understanding the transport mechanism of Fpn is to obtain sufficient quantities of pure and stable protein for downstream studies. As such, we detail here an expression and purification protocol for mouse Fpn yielding milligram quantities of pure protein. We have generated deletion constructs exhibiting enhanced thermal stability and which retained iron‐transport activity and hepcidin responsiveness, providing a platform for further biophysical studies of Fpn.

中文翻译：

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小鼠铁转运蛋白的分离和热稳定性

Ferroportin (Fpn) 是一种重要的哺乳动物铁转运蛋白，受激素铁调素的负调控。由于缺乏生化、生物物理和高分辨率结构研究，我们目前对 Fpn 介导的铁流出和调节的分子理解有限。了解 Fp​​n 转运机制的关键一步是获得足够数量的纯且稳定的蛋白质用于下游研究。因此，我们在这里详细介绍了小鼠 Fpn 的表达和纯化方案，可产生毫克量的纯蛋白质。我们已经生成了具有增强热稳定性并保留了铁转运活性和铁调素反应性的缺失构建体，为 Fpn 的进一步生物物理研究提供了平台。