title: Synthetic Lipid-Containing Scaffolds Enhance Production by Colocalizing Enzymes.
abstract: Subcellular organization is critical for isolating, concentrating, and protecting biological activities. Natural subcellular organization is often achieved using colocalization of proteins on scaffold molecules, thereby enhancing metabolic fluxes and enabling coregulation. Synthetic scaffolds extend these benefits to new biological processes and are typically constructed from proteins or nucleic acids. To expand the range of available building materials, we use a minimal set of components from the lipid-encapsulated bacteriophage ϕ6 to form synthetic lipid-containing scaffolds (SLSs) in E. coli. Analysis of diffusive behavior by particle tracking in live cells indicates that SLSs are >20 nm in diameter; furthermore, density measurements demonstrate that SLSs contain a mixture of lipids and proteins. The fluorescent proteins mCitrine and mCerulean can be colocalized to SLSs. To test for effects on enzymatic production, we localized two enzymes involved in indigo biosynthesis to SLSs. We observed a scaffold-dependent increase in indigo production, showing that SLSs can enhance the production of a commercially relevant metabolite.
authors.affiliation: Department of Systems Biology, Harvard Medical School , 02115, Boston, Massachusetts United States.|Wyss Institute for Biologically Inspired Engineering, Harvard University , 02115, Boston, Massachusetts United States.
authors.forename: Cameron
authors.initials: C
authors.lastname: Myhrvold
authors.affiliation: Department of Systems Biology, Harvard Medical School , 02115, Boston, Massachusetts United States.|Wyss Institute for Biologically Inspired Engineering, Harvard University , 02115, Boston, Massachusetts United States.
authors.forename: Jessica K
authors.initials: JK
authors.lastname: Polka
authors.affiliation: Department of Systems Biology, Harvard Medical School , 02115, Boston, Massachusetts United States.|Wyss Institute for Biologically Inspired Engineering, Harvard University , 02115, Boston, Massachusetts United States.
authors.forename: Pamela A
authors.initials: PA
authors.lastname: Silver
mesh_terms: D017126:Bacteriophage phi 6 / Q000737:chemistry*; D018914:Cells, Immobilized; D004926:Escherichia coli / Q000201:enzymology; D008055:Lipids / Q000737:chemistry*; D009696:Nucleic Acids / Q000737:chemistry; D014176:Protein Biosynthesis; D011506:Proteins / Q000737:chemistry*; D058615:Synthetic Biology
chemical_list: D008055:Lipids; D009696:Nucleic Acids; D011506:Proteins
keywords: bacteriophage ϕ6; metabolic engineering; protein fusions; scaffolds; subcellular organization