(48f) Engineering Synthetic Organelles: Encapsulating Heterologous Proteins Into Bacterial Microcompartments

Subcellular bacterial microcompartments are protein scaffolds of approximately 100 nm diameter that serve as a promising starting point for the creation of organelles to spatially organize biochemical reactions within the bacterial cell. Targeting enzymes or pathways of enzymes to such organelles would enable a wide variety of biotechnological applications by enhancing synthetic metabolic pathway yields, shielding the cell from toxic intermediates and products, and providing a distinct local environment for biochemical reactions that are incompatible with the cytosol. Here, we demonstrate the ability to encapsulate heterologous proteins into the lumen of bacterial microcompartments.  We then investigate the role of N-terminal signal sequence peptides in localizing proteins to microcompartments.  Through single amino acid substitution mutations, we find that a motif with specific spacing between charged and hydrophobic amino acids play an important role in making proteins competent for microcompartment localization.  These results will aid in the creation of a consensus sequence, and allow for the eventual colocalization of multiple enzymes to bacterial microcompartments to serve as nanobioreactors for biochemical pathways.