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Team:Minnesota/Project
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Overall project
Metabolic Engineering: In vivo Nanobioreactors
Modern microbial engineering methods allow the introduction of useful exogenous metabolic pathways into cells. Metabolism of certain organic compounds is sometimes limited by the production of toxic intermediates. Several bacteria have evolved protein based microcompartments capable of sequestering such reactions, thus protecting cytosolic machinery and processes from interference by these intermediates. For our project, we will identify and transform the genes encoding proteins responsible for the production and assembly of bacterial microcompartments. Additionally, we will confirm the signal sequences that target enzymes to the protein compartments by fusing this sequence to reporter genes. To demonstrate the microcompartment’s potential to serve as nanobioreactors, we will target genes encoding a short catabolic pathway into recombinant microcompartments assembled in E. coli.
Project Details: In vivo Nanobioreactors
For application in synthetic biology, bacterial microcompartments could be useful tools for engineering metabolic pathways for two important reasons. First, they can be used to sequester a toxic intermediate away from the cytoplasm. Second, by co-localizing enzymes inside a microcompartment the rate of diffusion of intermediate between enzymatic steps is decreased, thereby improving efficiency.
Background
A bacterial microcompartment (BMC) is a polyhedral protein complex that encapsulates enzymes involved in specific metabolic pathways. BMCs were first observed by microscope in cyanobacteria the 1950’s. By the late 1990’s, it had become clear that these proteinaceous structures are produced by many types of bacteria, and serve various functions (yeates et al. 2008). There are multiple examples of BMC’s occurring in nature: the carboxysome found in cyanobacteria and chemoautotrophic bacteria, the propanediol utilization compartment found in enterobacteria, as well as the ethanolamine utilization (Eut) compartment used in this project, also found in enterobacteria. By sequestering enzymes and substrates together in an enclosed space within the cell, the overall reaction efficiency is greatly improved. In some species, BMCs localize toxic compounds with enzymes that convert them into benign compounds. In this way a bacterial cell can ensure that the problematic intermediate is not released into the cell cytoplasm. BMCs are ideal for iGEM because they could be harnessed as nanobioreactors for metabolic engineering. Engineering a strain to express a BMC shell is possible because the genes that encode the BMC associated proteins are frequently organized tightly together into an operon. For this reason, theses are fairly well characterized. Engineering a BMC to house an enzymatic pathway would increase the efficiency of the process, and would also quarantine any toxic intermediates (Fig 1).
