title: Employing bacterial microcompartment technology to engineer a shell-free enzyme-aggregate for enhanced 1,2-propanediol production in Escherichia coli.
abstract: Bacterial microcompartments (BMCs) enhance the breakdown of metabolites such as 1,2-propanediol (1,2-PD) to propionic acid. The encapsulation of proteins within the BMC is mediated by the presence of targeting sequences. In an attempt to redesign the Pdu BMC into a 1,2-PD synthesising factory using glycerol as the starting material we added N-terminal targeting peptides to glycerol dehydrogenase, dihydroxyacetone kinase, methylglyoxal synthase and 1,2-propanediol oxidoreductase to allow their inclusion into an empty BMC. 1,2-PD producing strains containing the fused enzymes exhibit a 245% increase in product formation in comparison to un-tagged enzymes, irrespective of the presence of BMCs. Tagging of enzymes with targeting peptides results in the formation of dense protein aggregates within the cell that are shown by immuno-labelling to contain the vast majority of tagged proteins. It can therefore be concluded that these protein inclusions are metabolically active and facilitate the significant increase in product formation.
authors.affiliation: School of Biosciences, University of Kent, Giles Lane, Canterbury, Kent CT2 7NJ, UK.
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Lee
authors.affiliation: School of Biosciences, University of Kent, Giles Lane, Canterbury, Kent CT2 7NJ, UK.
authors.forename: Ian R
authors.initials: IR
authors.lastname: Brown
authors.affiliation: School of Biosciences, University of Kent, Giles Lane, Canterbury, Kent CT2 7NJ, UK.
authors.forename: Rokas
authors.initials: R
authors.lastname: Juodeikis
authors.affiliation: School of Biosciences, University of Kent, Giles Lane, Canterbury, Kent CT2 7NJ, UK. Electronic address: S.Frank@kent.ac.uk.
authors.forename: Stefanie
authors.initials: S
authors.lastname: Frank
authors.affiliation: School of Biosciences, University of Kent, Giles Lane, Canterbury, Kent CT2 7NJ, UK. Electronic address: m.j.warren@kent.ac.uk.
authors.forename: Martin J
authors.initials: MJ
authors.lastname: Warren
mesh_terms: D058045:Artificial Cells / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D053898:Biosynthetic Pathways / Q000502:physiology*; D002451:Cell Compartmentation / Q000502:physiology*; D004926:Escherichia coli / Q000502:physiology*; D024861:Genetic Enhancement / Q000379:methods*; D060847:Metabolic Engineering / Q000379:methods*; D053858:Metabolic Networks and Pathways / Q000502:physiology; D009097:Multienzyme Complexes / Q000235:genetics* / Q000378:metabolism; D019946:Propylene Glycol / Q000302:isolation & purification / Q000378:metabolism; D011506:Proteins / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D009097:Multienzyme Complexes; D011506:Proteins; D011993:Recombinant Fusion Proteins; D019946:Propylene Glycol
keywords: Biotechnology; Compartmentalisation; Metabolic engineering; Protein aggregation; Synthetic biology