Quantitative Description of Intrinsically Disordered Proteins Using Single-Molecule FRET, NMR, and SAXS,Journal of the American Chemical Society

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Quantitative Description of Intrinsically Disordered Proteins Using Single-Molecule FRET, NMR, and SAXS
Journal of the American Chemical Society ( IF 14.4 ) Pub Date : 2021-11-24 , DOI: 10.1021/jacs.1c06264
Samuel Naudi-Fabra ₁ , Maud Tengo ₁ , Malene Ringkjøbing Jensen ₁ , Martin Blackledge ₁ , Sigrid Milles ₁

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Studying the conformational landscape of intrinsically disordered and partially folded proteins is challenging and only accessible to a few solution state techniques, such as nuclear magnetic resonance (NMR), small-angle scattering techniques, and single-molecule Förster resonance energy transfer (smFRET). While each of the techniques is sensitive to different properties of the disordered chain, such as local structural propensities, overall dimension, or intermediate- and long-range contacts, conformational ensembles describing intrinsically disordered proteins (IDPs) accurately should ideally respect all of these properties. Here we develop an integrated approach using a large set of FRET efficiencies and fluorescence lifetimes, NMR chemical shifts, and paramagnetic relaxation enhancements (PREs), as well as small-angle X-ray scattering (SAXS) to derive quantitative conformational ensembles in agreement with all parameters. Our approach is tested using simulated data (five sets of PREs and 15 FRET efficiencies) and validated experimentally on the example of the disordered domain of measles virus phosphoprotein, providing new insights into the conformational landscape of this viral protein that comprises transient structural elements and is more compact than an unfolded chain throughout its length. Rigorous cross-validation using FRET efficiencies, fluorescence lifetimes, and SAXS demonstrates the predictive nature of the calculated conformational ensembles and underlines the potential of this strategy in integrative dynamic structural biology.

中文翻译：

使用单分子 FRET、NMR 和 SAXS 定量描述本质无序蛋白质

研究本质上无序和部分折叠的蛋白质的构象景观具有挑战性，并且只能通过几种溶液态技术来实现，例如核磁共振 (NMR)、小角度散射技术和单分子福斯特共振能量转移 (smFRET)。虽然每种技术对无序链的不同特性都很敏感，例如局部结构倾向、整体尺寸或中长程接触，但准确描述本质无序蛋白质 (IDP) 的构象整体应该理想地尊重所有这些特性。在这里，我们开发了一种集成方法，使用大量 FRET 效率和荧光寿命、NMR 化学位移和顺磁弛豫增强 (PRE) 以及小角 X 射线散射 (SAXS) 来导出与以下一致的定量构象系综：所有参数。我们的方法使用模拟数据（五组 PRE 和 15 个 FRET 效率）进行了测试，并以麻疹病毒磷蛋白无序结构域为例进行了实验验证，为这种病毒蛋白的构象景观提供了新的见解，该病毒蛋白包含瞬时结构元件，并且是比整个长度上展开的链条更紧凑。使用 FRET 效率、荧光寿命和 SAXS 进行严格的交叉验证，证明了计算的构象集合的预测性质，并强调了该策略在综合动态结构生物学中的潜力。

更新日期：2021-12-08

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