Department of Chemistry, Case Western Reserve University, Cleveland, Ohio, USA.
Roy J. Carver Department of Biochemistry, Biophysics and Molecular Biology, Iowa State University, Ames, Iowa, USA.
Proteins. 2021 Jul;89(7):781-791. doi: 10.1002/prot.26058. Epub 2021 Feb 15.
We have investigated the pressure- and temperature-induced conformational changes associated with the low complexity domain of hnRNP A1, an RNA-binding protein able to phase separate in response to cellular stress. Solution NMR spectra of the hnRNP A1 low-complexity domain fused with protein-G B1 domain were collected from 1 to 2500 bar and from 268 to 290 K. While the GB1 domain shows the typical pressure-induced and cold temperature-induced unfolding expected for small globular domains, the low-complexity domain of hnRNP A1 exhibits unusual pressure and temperature dependences. We observed that the low-complexity domain is pressure sensitive, undergoing a major conformational transition within the prescribed pressure range. Remarkably, this transition has the inverse temperature dependence of a typical folding-unfolding transition. Our results suggest the presence of a low-lying extended and fully solvated state(s) of the low-complexity domain that may play a role in phase separation. This study highlights the exquisite sensitivity of solution NMR spectroscopy to observe subtle conformational changes and illustrates how pressure perturbation can be used to determine the properties of metastable conformational ensembles.
我们研究了 hnRNP A1 低复杂度结构域与压力和温度诱导的构象变化之间的关系，hnRNP A1 是一种能够响应细胞应激而发生液-液相分离的 RNA 结合蛋白。我们从 1 巴到 2500 巴和从 268 开尔文到 290 开尔文收集了与蛋白-G B1 结构域融合的 hnRNP A1 低复杂度结构域的溶液 NMR 谱。虽然 GB1 结构域表现出小球形结构域中典型的压力诱导和冷温度诱导的解折叠，但 hnRNP A1 的低复杂度结构域表现出不寻常的压力和温度依赖性。我们观察到低复杂度结构域对压力敏感，在规定的压力范围内发生主要的构象转变。值得注意的是，这种转变具有典型折叠-去折叠转变的逆温度依赖性。我们的结果表明，低复杂度结构域存在一个低势的扩展和完全溶剂化的状态，这可能在液-液相分离中发挥作用。本研究突出了溶液 NMR 光谱学对观察细微构象变化的敏感性，并说明了压力扰动如何用于确定亚稳构象集合的性质。
