User:Paul Muir/Notebook/20.109(F11): Paul Muir and Cuong Nguyen

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Initial Ideas

Optogenetics:

Researchers can use genetically encoded light responsive proteins to control cellular pathways. This enables researchers to more easily alter cellular behavior in order to study various mechanisms. Some possible uses for light responsive proteins include light dependent allosteric inhibition of certain proteins in a pathway and light inducible protein-protein interactions in order to create light inducible localization of proteins {13 Jared,E.Toettcher 2011;}.


Metabolic Engineering:

Microbes can be engineered to take up starting materials and assemble a desired chemical product. This process involves importing the DNA encoding the various enzymes necessary for product formation. Microbial material production can lead to reductions in cost and in the use of hazardous chemicals. While this method shows great promise there are also issues surrounding integrating the reengineered pathway with the microbe’s natural metabolic pathways. If a balance between the two isn’t maintained then the cell could be starved of nutrients and die or no product could be formed. Careful manipulation of the promoters driving the engineered pathway and isolation of the engineered pathway’s enzymes in synthetic organelles is a possible solution to competition between the metabolic pathways {14 Keasling, Jay 2010;}.

Research Proposal

Optogenetics provides researchers with the ability to control cellular events on an almost instantaneous timescale. A common problem in metabolic engineering is the regulation of non-native metabolic pathways in the host organism. Through the use of light sensitive activation of enzymes and the light sensitive localization of proteins involved in a metabolic pathway a metabolic pathway can be more tightly regulated and synchronized across an entire population of cells. Metabolic pathways for the synthesis of artemisinin and alkanes have been developed by Jay Keasling and George Church. One possible project could involve utilizing optogenetics to optimize the pathway to increase yield. Perhaps a more immediately applicable and testable project could involve placing a step in a metabolic pathway which yields toxic metabolites under optogenetic control and determining whether activating the step only at certain times leads to a decrease in cell death due to the toxic metabolites being present in the cell for a shorter amount of time.

A frontier in metabolic engineering involves the construction of complex molecules from multiple chemical subunits all constructed and brought together in the same cell. A possible experiment could involve placing the enzymes that construct the various subunits under optogenetic control for localization to different membranes. One could then localize the construction of the various subunits to different parts of the cell using light to induce the localization. Once a sufficient number of subunits has been constructed one could turn off the light and end the localization. This would allow the subunits to interact and form the more complex molecules. The localization of the subunit synthesis to different membranes would also help minimize crosstalk between the different synthetic pathways present in the same cell.

Resources

Jared, E. Toettcher, et al. "The Promise of Optogenetics in Cell Biology: Interrogating Molecular Circuits in Space and Time." Nat Methods 8.1 (2011): 35-8.

Keasling, Jay. "Manufacturing Molecules through Metabolic Engineering." Science 330.6009 (2010): 1355-8.


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