Session L46: Invited Session: Design Principles in Synthetic Biology

Enzyme clustering can induce metabolic channeling

Direct channeling of intermediates via a physical tunnel between enzyme active sites is an established mechanism to improve metabolic efficiency. In this talk, I will present a theoretical model that demonstrates that coclustering multiple enzymes into proximity can yield the full efficiency benefits of direct channeling. The model predicts the separation and size of coclusters that maximize metabolic efficiency, and this prediction is in agreement with the spacing between coclusters in yeast and mammalian cells. The model also predicts that enzyme agglomerates can regulate steady--state flux division at metabolic branch points: we experimentally test this prediction for a fundamental branch point in \textit{Escherichia coli}, and the results confirm that enzyme colocalization~within an agglomerate can accelerate the processing of a shared intermediate by one branch. Our studies establish a quantitative framework to understand coclustering-mediated metabolic channeling and its application to both efficiency improvement and metabolic regulation.   

Programming Biology for Health and Sustainability

Biology presents us with an array of design principles. ~From studies of both simple and more complex systems, we understand some of the fundamentals of how Nature works. We are interested in using the foundations of biology to engineer cells in a logical and predictable way to perform certain functions. By necessity, the predictable engineering of biology requires knowledge of quantitative behavior of individual cells and communities. ~By building and analyzing synthetic systems, we learn more about the fundamentals of biological design as well as engineer useful living devices with myriad applications. ~For example, we are interested in building cells that can perform specific tasks, such as remembering past events and thus acting as biological computers. ~Moreover, we design cells with predictable biological properties that serve as cell-based sensors, factories for generating useful commodities and improved centers for carbon fixation. We have recently engineered natural gut bacteria that can non-invasively report on the dynamics of the animal gut. ~In doing so, we have made new findings about how cells interact with and impact on their environment. These principles can be applied to problems of all natural environments.   

Synthetic networks in microbial communities

While bacteria are single celled organisms, they predominantly reside in structured communities known as biofilms. Cells in biofilms are encapsulated and protected by the extracellular matrix (ECM), which also confines cells in space. During biofilm development, microbial cells are organized in space and over time. Little is known regarding the processes that drive the spatio-temporal organization of microbial communities. Here I will present our latest efforts that utilize synthetic biology approaches to uncover the organizational principles that drive biofilm development. I will also discuss the possible implications of our recent findings in terms of the cost and benefit to biofilm cells.   

Metabolite Valves: Dynamic Control of Metabolic Flux for Pathway Engineering

Microbial strains have been successfully engineered to produce a wide variety of chemical compounds, several of which have been commercialized. As new products are targeted for biological synthesis, yield is frequently considered a primary driver towards determining feasibility. Theoretical yields can be calculated, establishing an upper limit on the potential conversion of starting substrates to target compounds. Such yields typically ignore loss of substrate to byproducts, with the assumption that competing reactions can be eliminated, usually by deleting the genes encoding the corresponding enzymes. However, when an enzyme encodes an essential gene, especially one involved in primary metabolism, deletion is not a viable option. Reducing gene expression in a static fashion is possible, but this solution ignores the metabolic demand needed for synthesis of the enzymes required for the desired pathway. We have developed Metabolite valves to address this challenge. The valves are designed to allow high flux through the essential enzyme during an initial period where growth is favored. Following an external perturbation, enzyme activity is then reduced, enabling a higher precursor pool to be diverted towards the pathway of interest. We have designed valves with control at both the transcriptional and post-translational levels. In both cases, key enzymes in glucose metabolism are regulated, and two different compounds are targeted for heterologous production. We have measured increased concentrations of intracellular metabolites once the valve is closed, and have demonstrated that these increased pools lead to increased product yields. These metabolite valves should prove broadly useful for dynamic control of metabolic flux, resulting in improvements in product yields.