Exploring the Impact of Backbone Flexibility on Folding Mechanisms of Protein Mimetics: Integrating Experiment and Simulation

有了这个奖项，化学学部的生命过程化学项目资助了匹兹堡大学的Seth Horne博士和Lillian Chong博士，以表征人工蛋白质类分子的折叠机制。蛋白质是复杂的分子机器，执行维持生命所必需的大部分功能，由称为氨基酸的分子构建块组合而成，排列成长链。自然界提供了20种不同的氨基酸分子，它们连接在一起的序列的变化导致蛋白质采用不同的三维折叠形状。这些折叠的形状反过来又决定了蛋白质的功能。近年来，研究人员已经证明，各种人工蛋白质骨架（其中一些天然氨基酸被大小大致相同的不同分子取代）可以折叠成复杂的蛋白质样形状。人工骨架在帮助理解蛋白质如何工作方面具有基本价值，而且与天然骨架相比，它还具有实际的好处。小蛋白目前被广泛用于疾病的诊断和治疗；然而，这种应用的一个重要限制是血液中丰富的酶的快速降解。在保持侧链完整的情况下改变主链，可以制造出一种分子，这种分子模仿天然蛋白质原型的折叠形状和生物功能，但在体内不易降解。设计具有可预测形状的人工主干的一个关键部分是了解它们如何折叠。本项目旨在通过紧密耦合实验和计算模型来阐明人工骨干折叠的分子机制。这项工作包括开发一个新的力场——描述天然和人工氨基酸之间相互作用的数学模型——用于研究的建模部分。力场将通过广泛使用的AMBER分子动力学模拟包作为开源传播。这个跨学科的合作项目为参与研究的研究生和本科生提供了宝贵的培训场地，并支持各种教育和推广活动，包括夏季本科生创意科学写作研讨会，有关科学研究和科学传播问题的客座讲座，以及在当地一所高中举行的为期三周的Python编程研讨会。创意科学写作工作坊由匹兹堡大学化学系、英语系、科学系的历史和哲学系设计并提供，为学生提供科学和写作的联合指导，最终在新闻媒体上发表一篇或多篇具有广泛读者群的论文。最近的一项关键进展推动了蛋白质模拟物结构复杂性的前沿，即发现主链连通性可以在不损害天然侧链序列指定的折叠的情况下发生实质性改变。与这种“异质骨架”蛋白质模拟物的结构信息相比，实际上对骨架改变如何影响动力学或折叠途径一无所知。解决这一知识缺口有可能揭示人工骨架折叠行为的新见解，有助于设计更有效的蛋白质模拟物，并为与天然蛋白质折叠相关的基本问题提供见解。该项目正在确定蛋白质骨干连接如何影响链动力学和折叠机制。本研究的中心假设是，实验生物物理分析和原子计算机模拟相结合，可以揭示主链组成、折叠结构、链动力学、溶剂化和折叠途径之间复杂的相互作用。该项目包括设计和参数化改进的AMBER力场，用于模拟涉及人工蛋白质样主干的折叠和分子识别事件，以及开发使用加权集合增强采样策略模拟蛋白质折叠过程的新协议。力场作为AMBER分子动力学包的一部分正在传播。新的模拟策略被用于模拟在PI实验室合成的相应蛋白质模拟物上进行的热力学，结构和动力学实验。这种独特的实验和计算研究相结合，提供了对天然蛋白质和人工蛋白质之间稳定性差异的分子起源的见解，人工骨架蛋白质模拟物的折叠行为，以及骨架预组织在蛋白质折叠中的基本作用。该项目为高中生、本科生和研究生提供最新蛋白质设计和建模的研究培训机会。匹兹堡大学（University of Pittsburgh）正在扩大本科生暑期创意科学项目（Creative Science Summer Program），该项目旨在培养学生在科学领域有价值的写作技能，以交流科学在社会中扮演的关键角色。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

With this award, the Chemistry of Life Processes Program in the Division of Chemistry is funding Dr. Seth Horne and Dr. Lillian Chong from the University of Pittsburgh to characterize folding mechanisms in artificial, protein-like molecules. Proteins, the intricate molecular machines that perform the majority of functions necessary to sustain life, are made up of combinations of molecular building blocks known as amino acids, arrayed in long chains. Nature provides twenty different amino acid molecules, and variations in the sequence in which they are chained together leads proteins to adopt diverse three-dimensional folded shapes. These folded shapes, in turn, dictate protein function. In recent years, researchers have shown that a variety of artificial protein backbones, where some of the natural amino acids are replaced by different molecules of approximately the same size, can fold to adopt complex protein-like shapes. Artificial backbones have fundamental value in helping understand how proteins work and also offer practical benefits over their natural counterparts. Small proteins currently find widespread use as agents for the diagnosis and treatment of disease; however, an important limitation to such application is rapid degradation by enzymes that are abundant in the bloodstream. Changing the backbone while keeping side chains intact can create a molecule that mimics the folded shape and biological function of a prototype natural protein but is less prone to degradation in the body. A crucial component in the design of artificial backbones with predictable shapes is understanding how they fold. This project seeks to elucidate the molecular mechanism by which artificial backbones fold through closely-coupled experiments and computational modeling. This work includes the development of a new force field---the mathematical model describing interactions among the natural and artificial amino acids---for the modeling component of the research. The force field will be disseminated as open source through the widely-used AMBER molecular dynamics simulation package. This interdisciplinary, collaborative project is providing a valuable training ground for graduate and undergraduate students participating in the research, and is supporting diverse educational and outreach activities, including a summer undergraduate creative science writing workshop, guest lectures on issues related to scientific research and science communication, and a three-week workshop on Python coding at a local high school. The creative science writing workshop is being designed and offered by the Chemistry, English, and the History and Philosophy of Science Departments at the University of Pittsburgh, and provides students with joint science and writing mentorship, culminating in submission of one or more capstone pieces for publication in news outlets with broad readership.A key recent advance that pushed the frontier of structural complexity possible in protein mimetics is the finding that backbone connectivity can be substantially altered without compromising the fold specified by a natural side-chain sequence. Compared to the growing body of structural information on such "heterogeneous-backbone" protein mimics, virtually nothing is known about how backbone alteration impacts dynamics or folding pathways. Addressing this gap in knowledge has the potential to reveal new insights into the folding behavior in artificial backbones, aid in the design of more effective protein mimics, and provide insights into fundamental issues related to natural protein folding. This project is determining how protein backbone connectivity influences chain dynamics and folding mechanisms. The research is guided by the central hypothesis that a combination of experimental biophysical analysis and atomistic computer simulations can reveal the complex interplay among backbone composition, folded structure, chain dynamics, solvation, and folding pathways. The project includes the design and parameterization of a modified AMBER force field for simulating folding and molecular recognition events involving artificial protein-like backbones, and the development of novel protocols for simulating protein folding processes using the weighted ensemble enhanced sampling strategy. The force field is being disseminated as part of the AMBER molecular dynamics package. The new simulation strategies are being used to model the thermodynamic, structural, and kinetics experiments performed on corresponding protein mimetics synthesized in the PI's lab. This unique combination of experimental and computational research is providing insights into the molecular origins of stability differences between natural proteins and artificial counterparts, folding behavior in artificial-backbone protein mimetics, and the fundamental role of backbone preorganization in protein folding. The project is providing research training opportunities for high school, undergraduate, and graduate students in state-of-the-art protein design and modeling. An undergraduate Creative Science Summer Program at the University of Pittsburgh is being expanded, enabling students to develop valuable writing skills in the sciences, to communicate the critical roles that science plays in society.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.