Coiled-coil Modules for Molecular Engineering and Synthetic Biology

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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.