Entanglement in the Structural Biology of Living Systems

生命系统结构生物学中的纠缠

• 批准号: 2210636
• 负责人: Patricia Jennings
• 金额: $ 90万
• 依托单位: University of California-San Diego
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2027-08-31
• 项目状态: 未结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2210636&HistoricalAwards=false
• 关键词: Entanglement; Structural; Biology; Living; Systems

Protein folding from a linear polymer of amino acids into a three-dimensional shape is an essential process for life. The polymer may become hopelessly tangled and lead to loss of function and removal from the cell. However, entanglement in specific locations within the biopolymeric protein can lead to enhancement of stability, but the process by which specific entanglement occurs is not understood at the molecular level. This project seeks to address how a protein can form a deep intricate trefoil knot within the restrictive space of the cell, where proteins are formed, as well as what is the overall function of entanglement within the protein structure. Given the challenge and complexity of knotting a “string of pearls” into an enzyme capable of catalyzing a chemical reaction, the group proposes to combine the many facets of physical chemistry and physics to elucidate the unique yet critical role the knot plays in biology. Since the PI's discovery of a class of entangled pierced lasso proteins, this growing class now includes well over 350 protein families, spanning a huge range of fold types and biological functions. Through a combination of computational and physical experiments, the PI seeks to further the understanding of threading and knotting in proteins and expand upon their respective roles in biological function. These principles continue to be challenging topics in mathematics, physics and biology. Thus the continued investigation into the folding function interplay with the breadth of this study advances the concept of interplay in behavior and the strength of the interdisciplinary strategy. In the course of this project the PI's group will train undergraduate students in a variety of scientific areas related to knot formation and introduce them to the excitement of scientific inquiry. In addition, the PI has recruited students from diverse fields and (dis)abiledness to join the team of investigators, enriching the learning experience for all. Unexpectedly, proteins can tie themselves into knots in their native folds. Since protein folding from an unstructured polypeptide chain into the native-state is already complex, the existence of knotted polypeptide chains immediately raises the questions: how does the chain cross itself to form a knot and how does the knot affect function? To address these questions, the PI will study the deep-knotted trefoil methyltransferase (MT) families. Combining their strengths in this experimental and theoretical efforts, the PI’s group will expand their efforts in probing the mechanisms regulating knot threading/unthreading as well as the role of knots in the native state to ask how specific regions (identified as staples) within knots impact the fold, dynamics, and functions of proteins. Furthermore, the investigators will study whether specific regions identified as barriers to untying in the trefoil knotted proteins also contribute to functional regulation. Further exploration building upon these foundational studies allows them to investigate the interplay between the strained topology inherent in a knotted conformation and communication between functional sites. Their previous discoveries led to a protocol to explore threading mechanisms and untie a knotted protein. This experimental advantage allows the investigators to ask fundamental questions in the protein theory field: How can a protein form a threaded element? How does knotting affect function? How can it fold into a deeply knotted biologically active protein? To fully understand how nature controls the topology of proteins, extensive theoretical, numerical, and experimental progress is required. The goal of this project is to further expand upon these proof-of-principle experiments to explore both the structural and functional scope of the approach through an “all hands on deck” biochemical approach that will include molecular biology, protein chemistry, optical spectroscopy, and mass spectroscopy. This project requires the coordinated effort of top researchers working at the interface of mathematics, biology, chemistry, and physics. Ultimately, understanding the biophysical biochemistry of knotted proteins can be used to design and engineer novel classes of proteins that have superior mechanical and thermal stability properties. Knotting, threading and slip-knotting in proteins are new and challenging topics in knot theory for which new formalisms must be derived and must take into account physical properties of these biological objects.Broader impacts of this project include mentoring of K-12, and undergraduate and graduate students at UCSD. The PI has been involved in K-12 outreach activities by performing scientific experiments at local grammar schools, which are greater than 50% minority students, by performing experiments in special education schools, by helping to train children in middle school for the Science Olympiad, hosting High School students as research interns for the summer, and hosting people interested in a career in teaching in primary and secondary schools as research assistants in her laboratory. This project is funded by the Physics of Living Systems in the Division of Physics with support from the Molecular Biophysics Cluster in the Division of Molecular and Cellular Biosciences.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.

蛋白质从氨基酸的线性聚合物折叠成三维形状是生命的基本过程。聚合物可能变得不可救药地纠缠在一起，导致功能丧失和从细胞中移除。然而，生物聚合蛋白中特定位置的缠结可以导致稳定性的增强，但是在分子水平上，特定缠结发生的过程尚不清楚。该项目旨在解决蛋白质如何在细胞的限制性空间内形成深层复杂的三叶草结，以及蛋白质结构中纠缠的总体功能。考虑到将“珍珠链”结成一种能够催化化学反应的酶的挑战和复杂性，该小组建议将物理化学和物理学的许多方面结合起来，以阐明结在生物学中发挥的独特而关键的作用。自从PI发现一类缠绕的刺穿套索蛋白以来，这个不断增长的类别现在包括350多个蛋白质家族，涵盖了大量的折叠类型和生物功能。通过计算和物理实验的结合，PI试图进一步了解蛋白质中的穿线和打结，并扩展它们在生物功能中的各自作用。这些原理仍然是数学、物理和生物学中具有挑战性的话题。因此，对折叠功能相互作用的持续研究与本研究的广度推进了行为相互作用的概念和跨学科策略的力量。在这个项目的过程中，PI的小组将在与结形成相关的各种科学领域培养本科生，并将他们引入科学探究的兴奋。此外，PI还招募了来自不同领域和（残疾）能力的学生加入调查团队，丰富了所有人的学习经验。出乎意料的是，蛋白质可以把自己打成天然褶皱的结。由于蛋白质从非结构多肽链折叠到天然状态已经很复杂，打结多肽链的存在立即提出了问题：链如何交叉形成结，结如何影响功能？为了解决这些问题，PI将研究深结三叶甲基转移酶（MT）家族。结合他们在实验和理论方面的优势，PI的团队将扩大他们在探索调节结穿/脱线的机制以及结在原生状态下的作用方面的努力，以了解结内的特定区域（被确定为订钉）如何影响蛋白质的折叠、动力学和功能。此外，研究人员将研究三叶结蛋白中被确定为解结障碍的特定区域是否也有助于功能调节。在这些基础研究的基础上，进一步的探索使他们能够研究结构象中固有的应变拓扑与功能位点之间的通信之间的相互作用。他们之前的发现导致了一项探索穿线机制和解开打结蛋白的协议。这种实验优势使研究人员能够提出蛋白质理论领域的基本问题：蛋白质如何形成螺纹元素？打结如何影响功能？它是如何折叠成一个深结的生物活性蛋白质的？为了充分理解自然如何控制蛋白质的拓扑结构，需要广泛的理论、数值和实验进展。该项目的目标是进一步扩展这些原理验证实验，通过包括分子生物学、蛋白质化学、光谱学和质谱学在内的“全体人员参与”生化方法，探索该方法的结构和功能范围。这个项目需要顶尖的研究人员在数学、生物学、化学和物理学的交叉领域协同努力。最终，了解结蛋白的生物物理生化可以用于设计和制造具有优越机械和热稳定性的新型蛋白质。蛋白质中的结、线结和滑结是结理论中新的和具有挑战性的主题，必须推导出新的形式，并且必须考虑到这些生物对象的物理特性。该项目更广泛的影响包括指导加州大学圣地亚哥分校的K-12，本科生和研究生。PI参与了K-12的外展活动，在当地文法学校进行科学实验，这些学校有50%以上的少数民族学生，在特殊教育学校进行实验，帮助培训中学生参加科学奥林匹克竞赛，接待高中生作为暑期研究实习生，并接待对中小学教学事业感兴趣的人作为她实验室的研究助理。本项目由物理系生命系统物理系资助，并得到分子与细胞生物科学系分子生物物理集群的支持。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。

项目成果

Knotting Optimization and Folding Pathways of a Go-Model with a Deep Knot

深结 Go 模型的结优化和折叠路径

• DOI: 10.1021/acs.jpcb.2c05588
• 发表时间: 2022
• 期刊: The Journal of Physical Chemistry B
• 作者: Dahlstrom, Thomas J.;Capraro, Dominique T.;Jennings, Particia A.;Finke, John M.
• 通讯作者: Finke, John M.