From the Center for BioImaging Sciences and.
the Department of Biological Sciences, National University of Singapore, 14 Science Dr. 4, Singapore 117543, Singapore.
J Biol Chem. 2019 Apr 19;294(16):6468-6482. doi: 10.1074/jbc.RA118.007163. Epub 2019 Feb 28.
Dehydrins are intrinsically disordered proteins, generally expressed in plants as a response to embryogenesis and water-related stress. Their suggested functions are in membrane stabilization and cell protection. All dehydrins contain at least one copy of the highly conserved K-segment, proposed to be a membrane-binding motif. The dehydrin Lti30 () is up-regulated during cold and drought stress conditions and comprises six K-segments, each with two adjacent histidines. Lti30 interacts with the membrane electrostatically via pH-dependent protonation of the histidines. In this work, we seek a molecular understanding of the membrane interaction mechanism of Lti30 by determining the diffusion and molecular organization of Lti30 on model membrane systems by imaging total internal reflection- fluorescence correlation spectroscopy (ITIR-FCS) and molecular dynamics (MD) simulations. The dependence of the diffusion coefficient explored by ITIR-FCS together with MD simulations yields insights into Lti30 binding, domain partitioning, and aggregation. The effect of Lti30 on membrane lipid diffusion was studied on fluorescently labeled supported lipid bilayers of different lipid compositions at mechanistically important pH conditions. In parallel, we compared the mode of diffusion for short individual K-segment peptides. The results indicate that Lti30 binds the lipid bilayer via electrostatics, which restricts the mobility of lipids and bound protein molecules. At low pH, Lti30 binding induced lipid microdomain formation as well as protein aggregation, which could be correlated with one another. Moreover, at physiological pH, Lti30 forms nanoscale aggregates when proximal to the membrane suggesting that Lti30 may protect the cell by "cross-linking" the membrane lipids.
脱水蛋白是一种无序的内在蛋白，通常作为植物对胚胎发生和水相关压力的响应而表达。它们的建议功能是在膜稳定和细胞保护。所有脱水蛋白都至少包含一个高度保守的 K 段，被提议作为膜结合基序。脱水蛋白 Lti30（）在冷和干旱胁迫条件下上调，并包含六个 K 段，每个 K 段都有两个相邻的组氨酸。Lti30 通过组氨酸的 pH 依赖性质子化与膜静电相互作用。在这项工作中，我们通过确定 Lti30 在模型膜系统上的扩散和分子组织，通过全内反射荧光相关光谱（ITIR-FCS）和分子动力学（MD）模拟，寻求对 Lti30 膜相互作用机制的分子理解。通过 ITIR-FCS 探索扩散系数的依赖性以及 MD 模拟提供了对 Lti30 结合、结构域划分和聚集的深入了解。在机制上重要的 pH 条件下，研究了 Lti30 对不同脂质组成的荧光标记的支持脂双层上的膜脂扩散的影响。同时，我们比较了短的单个 K 段肽的扩散模式。结果表明，Lti30 通过静电相互作用结合脂质双层，从而限制了脂质和结合蛋白分子的流动性。在低 pH 下，Lti30 结合诱导脂质微区形成以及蛋白质聚集，这两者可以相互关联。此外，在生理 pH 下，当靠近膜时，Lti30 形成纳米级聚集体，这表明 Lti30 可能通过“交联”膜脂来保护细胞。
