FET: Small: Tools and Experimental Validation for Predicting Enzymatic Promiscuity and its Products

Tufts University, Medford MA

Investigators

Abstract

The engineering of microbial cells through synthetic biology promises to advance the production of high-volume commodity chemicals such as biopolymers, fuels, nutraceuticals, therapeutics, and other specialty products. Increasing understanding of metabolic and regulatory networks underlying microbial physiology hinges on developing metabolic models that capture enzymatic cellular activity. Despite significant progress in sequencing technology and model reconstruction, there are many cellular enzymatic activities that remain unknown. The hypothesis underlying this project is that undocumented promiscuous activity of enzymes results in the formation of unexpected reaction byproducts. This phenomenon is frequently observed by synthetic biologists, and sometimes exploited during design through ad hoc experimental efforts. However, there are currently limited ways to predict, analyze, or mitigate the effects of enzyme promiscuity. This project develops tools to predict stoichiometrically balanced reactions that reflect undocumented promiscuous enzymatic cellular activities. These reactions can be utilized to augment existing metabolic models and improve design tools. Further, the project experimentally validates the augmented models by engineering a microbial host to synthesize desirable chemicals and then analyzing host-pathway interactions. The proposed work is at the intersection of computer science and biology and will advance the ability to predict and assess the impact of enzyme promiscuity on biological systems. Training will be provided to graduate and undergraduate students and the research will be incorporated into classroom teaching. Underrepresented minorities in computing will be recruited to participate in the research through various established national and local programs. 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.