title: Diversion of flux toward sesquiterpene production in Saccharomyces cerevisiae by fusion of host and heterologous enzymes.
abstract: The ability to transfer metabolic pathways from the natural producer organisms to the well-characterized cell factory Saccharomyces cerevisiae is well documented. However, as many secondary metabolites are produced by collaborating enzymes assembled in complexes, metabolite production in yeast may be limited by the inability of the heterologous enzymes to collaborate with the native yeast enzymes. This may cause loss of intermediates by diffusion or degradation or due to conversion of the intermediate through competitive pathways. To bypass this problem, we have pursued a strategy in which key enzymes in the pathway are expressed as a physical fusion. As a model system, we have constructed several fusion protein variants in which farnesyl diphosphate synthase (FPPS) of yeast has been coupled to patchoulol synthase (PTS) of plant origin (Pogostemon cablin). Expression of the fusion proteins in S. cerevisiae increased the production of patchoulol, the main sesquiterpene produced by PTS, up to 2-fold. Moreover, we have demonstrated that the fusion strategy can be used in combination with traditional metabolic engineering to further increase the production of patchoulol. This simple test case of synthetic biology demonstrates that engineering the spatial organization of metabolic enzymes around a branch point has great potential for diverting flux toward a desired product.
authors.affiliation: Center for Microbial Biotechnology, Department of Systems Biology, Technical University of Denmark, Building 223, DK-2800 Kgs. Lyngby, Denmark.
authors.forename: Line
authors.initials: L
authors.lastname: Albertsen
authors.forename: Yun
authors.initials: Y
authors.lastname: Chen
authors.forename: Lars S
authors.initials: LS
authors.lastname: Bach
authors.forename: Stig
authors.initials: S
authors.lastname: Rattleff
authors.forename: Jerome
authors.initials: J
authors.lastname: Maury
authors.forename: Susanne
authors.initials: S
authors.lastname: Brix
authors.forename: Jens
authors.initials: J
authors.lastname: Nielsen
authors.forename: Uffe H
authors.initials: UH
authors.lastname: Mortensen
mesh_terms: D001709:Biotechnology / Q000379:methods; D051228:Geranyltranstransferase / Q000235:genetics / Q000378:metabolism*; D007535:Isomerases / Q000235:genetics / Q000378:metabolism*; D019686:Lamiaceae / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D015202:Protein Engineering / Q000379:methods; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D012717:Sesquiterpenes / Q000378:metabolism*
chemical_list: D010940:Plant Proteins; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D012717:Sesquiterpenes; D051228:Geranyltranstransferase; D007535:Isomerases; C065995:patchoulol synthase; C052983:patchouli alcohol