Collaborative Research: Multiscale molecular simulations of protein-mediated bilayer fusion

智力优势也许我们所知道的细胞生命最重要的结构是脂质双层。脂质分子由水溶性的“头”和水不溶性的“尾”组成，自发地组装成类脂质双层膜，其包围所有活细胞并进一步划分所有真核生物的细胞内部-植物、真菌、动物和人类所属的生命领域。典型的真核细胞中的膜网络非常复杂且高度动态：小隔间像气泡一样从某些膜上发芽，将货物从细胞的一部分运送到另一部分，在那里它们可以与其他膜融合，包括细胞的外膜。因此，双层融合是一种普遍存在的生物过程，与物质和信息的运输紧密相关，因此它受到几类膜相关蛋白的精细控制。这些蛋白质显然在融合膜上工作，但它们在分子尺度上诱导的复杂的几何和拓扑形状转换序列在实验中是不可能直接观察到的。相比之下，分子模拟为这些细节提供了一个窗口，但到目前为止，相关的长度和时间尺度已经被证明太大，以至于无法观察到现实系统大小的单个融合事件。该项目建立了两个研究人员之间的合作，旨在通过将多尺度粗粒度建模的最新进展与增强采样分子模拟相结合来应对这一挑战。由于这种策略允许结合重要的化学细节，同时代表大规模的膜变形，研究人员将能够阐明分子水平的机制如何在相关的生理长度和时间尺度上驱动融合事件。该项目通过三个阶段进行，即：（i）模拟原始双层与增强采样的融合，（ii）模型融合蛋白质的粗粒度模型的开发，SNARE系统，以及（iii）将这两个步骤结合成一个单一的方法。该项目将探讨许多与能量、形态和机械相关的主题，特别是围绕所谓的半融合中间态的问题，其中两个外层双层小叶已经融合，但由两个内层小叶形成的膜仍然将两个隔室分开。更广泛的影响由于双层融合在各种生物学中的重要性，该项目将影响生物科学中的许多主题。包括细胞内运输，病毒进入，神经递质释放，受精等生物过程。除了正在研究的具体问题之外，这里设想的计算方法采取了早期步骤，有效地模拟更复杂的多蛋白质/多膜现象，因此将有利于更广泛的一类分子生物学主题的未来研究。为了扩大研究成果的适用性，该项目中开发的模拟框架将免费提供教程，以支持有效的学习，并促进现有技术和模块向新应用的转变。该项目建立了工程和（生物）物理之间的跨学科交流，为该项目指导的学生的学术成长培养了一个刺激的跨学科环境。它将进一步将理论和计算方法从工程和物理学转移到生命科学及其日益量化的问题。双层融合的普遍存在及其与生物物理学中广泛的迷人主题的联系，这本身就是一个有趣的跨学科主题，也为该项目中开发的专业知识提供了极好的机会，以通过课堂材料，讲座演示，和公开讲座，两位研究人员将根据他们的经验和各自机构现有的成功项目实施这些活动。

INTELLECTUAL MERITPerhaps the most important structure for cellular life as we know it is the lipid bilayer. Lipid molecules, consisting of a water-soluble "head" and water-insoluble "tails", spontaneously assemble into sandwich-like bilayer membranes, which surround all living cells and further compartmentalize the cellular interiors of all eukaryotic organisms the domain of life to which plants, fungi, animals, and humans belong. The network of membranes in a typical eukaryotic cell is very complex and highly dynamic: small compartments bud off from certain membranes like bubbles, carrying cargo from one part of the cell to another, where they can fuse with yet other membranes, including the outer membrane of the cell. Bilayer fusion is therefore a ubiquitous biological process, tightly linked to the transport of material and information, and therefore it is exquisitely controlled by several classes of membrane-associated proteins. These proteins clearly perform work on the fusing membranes, but the intricate sequence of geometric and topological shape transformations they induce on the molecular scale are impossible to observe directly in experiment. In contrast, molecular simulation offers a window onto these details, but until now the relevant length- and time-scales have proven too big to observe even a single fusion event for a realistic system size. This project establishes a collaboration between two investigators with the aim to meet this challenge by combining recent advances in multiscale coarse-grained modeling with enhanced-sampling molecular simulation. Since this strategy allows incorporating important chemical detail while simultaneously representing large-scale membrane deformations, the investigators will be able to elucidate how molecular-level mechanisms drive fusion events across the relevant physiological length- and time-scales. The project proceeds through three phases, namely: (i) modeling the fusion of pristine bilayers with enhanced sampling, (ii) development of coarse-grained models of model fusogenic proteins, the SNARE system, and (iii) combining these two steps into a single methodology. The project will pursue many topics of energetic, morphological, and mechanistic relevance, in particular questions revolving around the so-called hemifusion intermediate state, for which the two outer bilayer leaflets have already fused but a membrane formed by the two inner leaflets still separates the two compartments.BROADER IMPACTSThis project will impact many topics in the biological sciences due to the central importance of bilayer fusion in a variety of biological processes, including intracellular trafficking, viral entry, neurotransmitter release, fertilization, and more. Beyond the specific questions under study, the computational approach envisioned here takes early steps towards efficient simulation of more complicated multiple-protein/multiple-membrane phenomena and will therefore benefit future studies of a wider class of molecular biological topics. To broaden applicability of the research outcomes, the simulation framework developed in this project will be made freely available with tutorials that will support efficient learning and facilitate the transformation of existing techniques and modules towards novel applications. This project establishes cross-disciplinary exchange between engineering and (bio)physics, fostering a stimulating interdisciplinary environment for the academic growth of students mentored in this project. It will further the transfer of theoretical and computational methodologies from engineering and physics into the life sciences and their increasingly quantitative set of problems. The ubiquity of bilayer fusion and its connection to a wide class of fascinating themes in biological physics, which is in itself an intriguing cross-disciplinary subject, also present excellent opportunities for the expertise developed in this project to feed outreach specifically tailored towards groups underrepresented in STEM fields for instance through classroom material, lecture demonstrations, and public talks and both investigators will implement such activities, building on both their experience and existing successful programs at their respective institutions.