Team:Virginia/Poster

Manifold: Protein Shells with Encapsulated DNA Scaffolds for Increasing Efficiency of Biosynthetic Pathways
Presented by Team Virginia 2020

Team: J. Ball, V. Gutierrez, C. Haws, A. Kola, S. Link, C. Marino, E. Micklovic, D. Patel, J. Polzin, A. Pradhan, P. Revelli

Advisors: K. G. Kozminski, J. A. Papin

Abstract:
The lack of a versatile and reliable way to improve metabolic flux channeling, pathway orthogonality, and product yields is a major impediment to the expanded utilization of biosynthesis for the production of drugs and industrially valuable chemicals. Manifold, a platform technology that addresses this problem, consists of bacterial microcompartments (BMCs) with encapsulated dsDNA scaffolds that sequester and spatially organize, at fixed concentrations, biosynthetic enzymes presented as zinc-finger fusion proteins. Here we deliver the designs for an E. coli cell capable of synthesizing resveratrol using the Manifold platform. The Manifold platform will help lower costs and expand the applications of chemical biosynthesis.
Inspiration
The field of synthetic biology has seen great success in utilizing bacteria and yeast to produce chemicals of medical and industrial value, but these methods still have the potential to be improved. In many cases, prokaryotes are structurally simpler and generally easier to engineer than eukaryotes, but do not possess a simple way to compartmentalize reaction pathways like eukaryotic organelles do. We strove to combine the ease of use of prokaryotes with the compartmentalization capability of eukaryotes by developing a method to synthetically isolate a simple biosynthetic pathway - resveratrol synthesis - within a prokaryotic system.
4 Big Problems in Biosynthesis
  • Flux imbalances occur when the amount of substrate available does not match the efficiency of the enzyme. In multi-enzyme pathways, this results in an overabundance or lack of intermediates, even with careful promoter and ribosome binding site choice.
  • Loss of intermediates can occur as they cross a membrane or move to a region of the cell where pathway enzymes are not present and thus, overall yields are lower than expected.
  • Pathway competition also reduces yield. If an enzyme, substrate, intermediate, or coenzyme of the pathway of interest is utilized by a process native to the cell it will be less likely to be available for the desired pathway.
  • Toxic intermediates can cause harm or cell death before any product can be made making biosynthesis highly inefficient or impossible for many pathways.
Problem
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Idea
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Section 1
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Section 2.1
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Section 2.2
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Section 3
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Results
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References and Acknowledgements
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