Dynamic nuclear polarization illuminates key protein-lipid interactions in the native bacterial cell envelope.
Dynamic nuclear polarization illuminates key protein-lipid interactions in the native bacterial cell envelope.
复制标题
动态核极化阐明了天然细菌细胞包膜中关键的蛋白质-脂质相互作用。
DOI：
10.1101/2023.05.18.541325
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发表时间：
2023
期刊：
影响因子：
--
通讯作者：
Marassi,FrancescaM
中科院分区：
文献类型：
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作者：
Kent,JamesE;Ackermann,BryceE;Debelouchina,GaliaT;Marassi,FrancescaM
阐明天然环境中蛋白质的结构和相互作用是结构生物学的基本目标。核磁共振 (NMR) 光谱非常适合这项任务，但通常灵敏度较低，尤其是在复杂的生物环境中。在这里，我们使用一种称为动态核极化（DNP）的灵敏度增强技术来克服这一挑战。我们应用 DNP 捕获外膜蛋白 Ail 的膜相互作用，外膜蛋白 Ail 是鼠疫耶尔森氏菌宿主入侵途径的关键组成部分。我们表明，天然细菌细胞包膜中 Ail 的 DNP 增强 NMR 谱得到了很好的解析，并且相关性丰富，而这在传统的固态 NMR 实验中是不可见的。此外，我们证明了 DNP 捕获蛋白质与周围脂多糖层之间难以捉摸的相互作用的能力。我们的结果支持一个模型，其中细胞外环精氨酸残基重塑膜环境，这一过程对于宿主入侵和发病机制至关重要。
Elucidating the structure and interactions of proteins in native environments is a fundamental goal of structural biology. Nuclear magnetic resonance (NMR) spectroscopy is well suited for this task but often suffers from low sensitivity, especially in complex biological settings. Here, we use a sensitivity-enhancement technique called dynamic nuclear polarization (DNP) to overcome this challenge. We apply DNP to capture the membrane interactions of the outer membrane protein Ail, a key component of the host invasion pathway ofYersinia pestis. We show that the DNP-enhanced NMR spectra of Ail in native bacterial cell envelopes are well resolved and enriched in correlations that are invisible in conventional solid-state NMR experiments. Furthermore, we demonstrate the ability of DNP to capture elusive interactions between the protein and the surrounding lipopolysaccharide layer. Our results support a model where the extracellular loop arginine residues remodel the membrane environment, a process that is crucial for host invasion and pathogenesis.