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Team:TorontoMaRSDiscovery/Project
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Engineering bacterial micro-compartments to investigate metabolic channeling and its potential uses in biotechnological applications
Background
It is well established that genes and their products do not operate in isolation but rather form parts of integrated biochemical pathways. There is increasing evidence that in many pathways, individual components exhibit varying degrees of spatial organization ranging from sub-cellular compartmentalization to the formation of discrete complexes.
For metabolic processes, the co-localization of enzymatic components has been shown to promote the transfer of substrates between consecutive reactions in a process termed “channeling”1. Metabolic channeling is defined as the process in which the intermediate produced by one enzyme is transferred to the next enzyme without complete mixing in the bulk phase12. Channeling results in the efficient translocation of substrates between enzymes and has been proposed to result in the following benefits: 1) Optimization of catalytic efficiency by decreasing transit time for intermediates; 2) Relief of the effects of product inhibition; 3) Protection from the creation of potentially toxic or unstable intermediates; and 4) Regulation of substrate flux through mediating pathway cross-talk 1. Potential applications range from the production of valuable compounds such as therapeutic molecules and biofuels2 to the degradation of toxic wastes3.
We propose a novel interdisciplinary project to systematically investigate how nature implements metabolic channeling and how this knowledge may be exploited for biotechnological applications.
Experimental Approach
Objectives
- Design, construct and characterize a micro-compartment expression system in E. coli.
- Demonstrate in vivo assembly of the expressed micro-compartments.
- Target a fluorescent marker (eCFP) to the micro-compartment.
- Identify and prioritize candidate enzyme pairs for channeling.
- Apply channeling to selected enzyme pairs.
