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Real-time tracking of protein unfolding with time-resolved x-ray solution scattering
Structural Dynamics ( IF 2.3 ) Pub Date : 2020-09-22 , DOI: 10.1063/4.0000013
L Henry 1 , M R Panman 1 , L Isaksson 1 , E Claesson 1 , I Kosheleva 2 , R Henning 2 , S Westenhoff 1 , O Berntsson
Affiliation  

The correct folding of proteins is of paramount importance for their function, and protein misfolding is believed to be the primary cause of a wide range of diseases. Protein folding has been investigated with time-averaged methods and time-resolved spectroscopy, but observing the structural dynamics of the unfolding process in real-time is challenging. Here, we demonstrate an approach to directly reveal the structural changes in the unfolding reaction. We use nano- to millisecond time-resolved x-ray solution scattering to probe the unfolding of apomyoglobin. The unfolding reaction was triggered using a temperature jump, which was induced by a nanosecond laser pulse. We demonstrate a new strategy to interpret time-resolved x-ray solution scattering data, which evaluates ensembles of structures obtained from molecular dynamics simulations. We find that apomyoglobin passes three states when unfolding, which we characterize as native, molten globule, and unfolded. The molten globule dominates the population under the conditions investigated herein, whereas native and unfolded structures primarily contribute before the laser jump and 30 μs after it, respectively. The molten globule retains much of the native structure but shows a dynamic pattern of inter-residue contacts. Our study demonstrates a new strategy to directly observe structural changes over the cause of the unfolding reaction, providing time- and spatially resolved atomic details of the folding mechanism of globular proteins.

中文翻译:

利用时间分辨 X 射线溶液散射实时跟踪蛋白质展开

蛋白质的正确折叠对其功能至关重要,蛋白质错误折叠被认为是多种疾病的主要原因。蛋白质折叠已经通过时间平均方法和时间分辨光谱进行了研究,但实时观察展开过程的结构动力学具有挑战性。在这里,我们演示了一种直接揭示展开反应中结构变化的方法。我们使用纳秒到毫秒时间分辨的 X 射线溶液散射来探测脱辅基肌红蛋白的展开。展开反应是通过纳秒激光脉冲引起的温度跳跃触发的。我们展示了一种解释时间分辨 X 射线溶液散射数据的新策略,该策略评估从分子动力学模拟获得的结构集合。我们发现脱辅基肌红蛋白在展开时会经历三种状态,我们将其描述为天然状态、熔球状态和展开状态。在本文研究的条件下,熔球在总体中占主导地位,而自然结构和未折叠结构分别主要在激光跳跃之前和 之后30 μs做出贡献。熔球保留了大部分天然结构,但显示出残留物间接触的动态模式。我们的研究展示了一种新策略,可以直接观察展开反应原因的结构变化,提供球状蛋白折叠机制的时间和空间解析原子细节。
更新日期:2020-10-30
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