Bacterial microcompartments are subcellular bacterial organelles bound by a porous protein membrane. Enteric and other bacteria use these organelles to sequester metabolic pathways to enhance growth and pathogenicity. We aim to develop a tunable nanobioreactor system based on these protein structures to spatially organize heterologous biosynthetic pathways in bacterial hosts. To this end, we develop tools to control both the formation of the microcompartments and their enzymatic content.
By engineering the transcriptional regulation of the microcompartments, we can orchestrate their formation and exercise control over the loading of heterologous enzymes to the nanobioreactor, tuning the catalytic activity of the microcompartments. We also engineer the N-terminal signal sequences responsible for enzyme targeting to the compartments, creating new signal sequences based on a common hydrophobic motif and allowing fine control over the loading phenotype of the reactors. Our results have implications for the controlled encapsulation of heterologous synthetic pathways to control enzyme stoichiometry and small molecule transport.