Coiled-coil Modules for Molecular Engineering and Synthetic Biology

智力优势：了解氨基酸序列如何决定蛋白质的结构和功能，为在纳米水平上设计分子工具提供了机会。螺旋螺旋是一种简单的蛋白质结构，由两条或更多的链组成，这些链连接成一个棒状的超级螺旋。几十年的研究已经深入了解了决定两条蛋白质链能否结合形成盘绕线圈的原理。这个过程现在可以控制，在一定程度上，通过操纵蛋白质序列使用合理的分子设计技术。该项目旨在利用当前对线圈序列-结构关系的理解，以及强大的分子工程技术，开发适用于构建新材料和使用线圈基序重新设计细胞电路的试剂。目标是生成一个完整描述的分子部分列表，以便其他科学家在生物工程项目中使用这些作为模块化单元。这项研究建立在先前鉴定的23种参与各种复杂相互作用模式的合成蛋白质的基础上。该项目的目标是：(1)测试这些先前在实验室溶液中表征的肽是否保持其在细菌和酵母内部的相互作用特性；(2)通过合理的分子设计来完善和调整这23种蛋白质的相互作用强度和特异性；(3)探索模块化卷曲线圈单元如何用于改变实验室酵母的信息处理，特别是通过修改map -激酶信号。(4)探索利用模块化卷曲蛋白构建人工dna结合蛋白。这些领域的成功将为合成线圈在分子工程应用中的广泛应用铺平道路。作为适应于自然界的模块化设计模块，线圈具有推进材料科学的潜力，为工程微生物细胞提供了新的能力，并与纳米级无机材料相结合，使先进的信息处理和智能设备成为可能。更广泛的影响：除了科学影响之外，该项目将为不同群体的年轻科学家提供综合教育/研究机会。拟议的工作将在麻省理工学院生物系的基廷实验室进行，那里的教育和研究活动是紧密结合的。基廷研究小组由研究生、博士后科学家和本科生组成，共有12人。具有不同科学、经济、种族和文化背景的实验室成员参与了所有的小组活动，基廷博士还参与了针对女性和科学领域代表性不足的学生的指导活动。本科生将被招募来参与这个项目，夏季邀请将特别扩展到那些不太可能接触到现代研究的个人。蛋白质工程和合成生物学是吸引本科生的研究课题，因为它们提供了在分子世界中合理构建全新事物的机会。这将增强探索和发现的精神，而这种精神对于吸引和留住年轻科学家至关重要。

Intellectual Merit: Understanding how amino-acid sequences determine the structure and function of proteins provides an opportunity to engineer molecular tools at the nanoscale level. The alpha helical coiled coil is a simple protein structure made up of two or more chains that associate into a rod-like super-helix. Several decades of research have provided insights into the principles that determine whether two protein chains can associate to form a coiled coil. This process can now be controlled, to a certain extent, by manipulating protein sequence using rational molecular design techniques. This project aims to exploit current understanding of coiled-coil sequence-structure relationships, along with powerful molecular engineering techniques, to develop reagents suitable for building new materials and re-engineering cellular circuits using coiled coil motifs. The goal is to generate a list of fully described molecular parts that will allow other scientists to use these as modular units in bioengineering projects. The research builds on the prior identification of 23 synthetic proteins that participate in a variety of complex interaction patterns. Goals for the project are to (1) test whether these peptides, previously characterized in solution in the laboratory, maintain their interaction properties inside bacteria and yeast, (2) refine and tune the interaction strengths and specificities of these 23 proteins through rational molecular design, (3) explore how the modular coiled-coil units can be used to alter information processing in laboratory yeast, specifically by modifying MAP-kinase signaling, and (4) explore the use of modular coiled-coil proteins for constructing artificial DNA-binding proteins. Success in these areas will pave the way for broad use of synthetic coiled coils in molecular engineering applications. As modular design blocks adapted from the natural world, coiled coils have the potential to advance materials science, provide new capabilities for engineering microbial cells, and interface with nanoscale inorganic materials, enabling advanced information processing and smart devices.Broader Impacts: Beyond the scientific impact, this project will enhance integrated educational/research opportunities for a diverse group of young scientists. The proposed work will be carried out in the Keating laboratory in the MIT biology department, where education and research activities are tightly coupled. The Keating research group of 12 individuals is composed of graduate students, postdoctoral scientists and undergraduate students. Laboratory members with diverse scientific, economic, ethnic and cultural backgrounds are involved in all group activities, and Dr. Keating additionally participates in mentoring activities for women and under-represented students in science. Undergraduate students will be recruited to work on this project, with summer invitations extended particularly to individuals not likely otherwise to be exposed to modern research. Protein engineering and synthetic biology are appealing research subjects for undergraduates, because these provide the opportunity to rationally construct something entirely new in the molecular world. This will enhance the spirit of exploration and discovery that is critical for engaging and retaining young scientists.