Combining Simulation and Spectroscopy to Determine the Structure and Dynamics of Adsorbed Proteins - Application to Biomass Conversion

在许多情况下，社会设计将可再生资源转化为有用产品的新系统的能力取决于界面过程（例如，酶转化不溶性多糖）。这些界面在原子尺度上的研究具有难以置信的挑战性，在过去十年中，在工程改进系统方面取得的成功相对较少。因此，我们建议华盛顿大学和马克斯普朗克聚合物研究所合作研究，将最先进的多尺度分子建模工具与和频率产生光谱（SFG）深度集成。酶促生物质转化，像许多生物过程一样，是由界面现象控制的速率。在生物质转化的情况下，不溶性纤维素微纤维造成扩散限制，因为反应不能在碰撞和适当的排列/结合之前进行。纤维素酶已经进化出精确的碳水化合物结合模块（CBMs），以增加界面附近酶活性位点的浓度，从而导致伴随的速率提高。一些CBMs还可以通过有益地改变微纤维结构来提高速率。然而，由于研究界面处蛋白质结构的普遍困难，对特异性CBM结合的结构基础知之甚少。我们的问题是：纤维素结合的结构动机是什么？哪些氨基酸侧链结合纤维素表面？本研究项目的成功完成将为界面生物催化提供大量新的基础知识和应用知识。具体来说，通过提供纤维素表面上结合模块的原子尺度结构和动力学，我们将为纤维素酶作用机制的控制动力学和能量贡献提供必要的见解。这些信息是提高催化速率的合理工程研究的先驱。更广泛地说，我们的分子模拟和SFG实验强耦合方法的成功演示将改变研究人员在界面上研究生物分子的方式。我们相信这项工作将导致研究界面生物催化的一般计算/实验框架，目前还不存在。

Society's ability to engineer new systems for converting renewable resources into useful products is, in many cases, dependent on interfacial processes (e.g., enzymatic conversion of an insoluble polysaccharide). These interfaces are incredibly challenging to study at the atomic scale, with comparatively little success over the last decade in engineering improved systems. Therefore, we propose a collaborative research effort between the University of Washington and the Max Planck Institute for Polymer Research to deeply integrate state of the art multiscale molecular modeling tools with sum frequency generation spectroscopy (SFG). Enzymatic biomass conversion, like many biological processes, is rate-controlled by interfacial phenomena. In the case of biomass conversion, the insoluble cellulose microfibril creates diffusion limitations as the reaction cannot proceed prior to collision and proper alignment/binding. Cellulase enzymes have evolved precise carbohydrate binding modules (CBMs) to provide an increase in concentration of enzyme active sites near the interface, which leads to concomitant rate enhancement. Some CBMs may also promote rates by beneficially altering the microfibril structure. However, due to the general difficulty of studying protein structure at interfaces, only very little is known about the structural basis of specific CBM binding. Questions we ask are: What structural motives are involved in cellulose binding? What amino acids side chains bind the cellulose surface? Successful completion of this research project will lead to a wealth of new fundamental and applied knowledge about interfacial biocatalysis. Specifically, by providing the atomic scale structure and dynamics of the binding module on a cellulose surface we will provide needed insight into the governing kinetic and energetic contributions that give rise to the mechanism of cellulase action. Such information is a precursor for rational engineering studies to improve catalytic rates. More broadly, successful demonstration of our approach to strongly couple molecular simulation and SFG experiments will transform the way researchers investigate biomolecules at interfaces. We believe this work will lead to a general computational/experimental framework for studying interfacial biocatalysis, which does not currently exist.