451591 Engineering the Shell Proteins of a Bacterial Microcompartment to Control Small Molecule Transport

Thursday, November 17, 2016: 12:30 PM
Continental 7 (Hilton San Francisco Union Square)
Marilyn F. Slininger1, Christopher Jakobson1 and Danielle Tullman-Ercek1,2, (1)Chemical and Biomolecular Engineering, University of California, Berkeley, Berkeley, CA, (2)Chemical and Biological Engineering, Northwestern University, Evanston, IL

Many non-native reactions in bacterial hosts have unacceptably low yields due to pathway toxicity or competition with other cellular processes. Sequestration of pathways within a diffusion barrier is a potential solution to these problems. Compartmentalization can prevent escape of volatile or toxic intermediates, prevent off-pathway reactions, and create private cofactor pools. Bacterial microcompartments are a class of proteinaceous organelles comprising a characteristic protein shell enclosing a set of enzymes. To enhance the function of heterologous pathways by encapsulation within synthetic microcompartments, we must understand how to control diffusion in and out of the microcompartment organelle. To this end, we explored how small differences in shell protein structure result in changes in the diffusion of metabolites through the shell by comparing the ethanolamine utilization (Eut) and the1,2-propanediol utilization (Pdu) microcompartments. We found that the Eut protein EutM properly incorporates into the Pdu microcompartment to form chimeric MCPs. This substitution alters native metabolite accumulation and the resulting growth on 1,2-propanediol. Further, we identified a single pore-lining residue mutation that confers the same phenotype as substitution of the full EutM protein, indicating that small molecule diffusion through the shell is the cause of growth enhancement. Finally, we show that the hydropathy index and charge of pore amino acids are important indicators to predict how pore mutations will affect growth on 1,2-propanediol, likely by controlling diffusion of one or more metabolites. Toggling pore properties will allow metabolic engineers to selectively control small molecule diffusion for the optimization of heterologous pathways.

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