title: Efficient diterpene production in yeast by engineering Erg20p into a geranylgeranyl diphosphate synthase.
abstract: Terpenes have numerous applications, ranging from pharmaceuticals to fragrances and biofuels. With increasing interest in producing terpenes sustainably and economically, there has been significant progress in recent years in developing methods for their production in microorganisms. In Saccharomyces cerevisiae, production of the 20-carbon diterpenes has so far proven to be significantly less efficient than production of their 15-carbon sesquiterpene counterparts. In this report, we identify the modular structure of geranylgeranyl diphosphate synthesis in yeast to be a major limitation in diterpene yields, and we engineer the yeast farnesyl diphosphate synthase Erg20p to produce geranylgeranyl diphosphate. Using a combination of protein and genetic engineering, we achieve significant improvements in the production of sclareol and several other isoprenoids, including cis-abienol, abietadiene and β-carotene. We also report the development of yeast strains carrying the engineered Erg20p, which support efficient isoprenoid production and can be used as a dedicated chassis for diterpene production or biosynthetic pathway elucidation. The design developed here can be applied to the production of any GGPP-derived isoprenoid and is compatible with other yeast terpene production platforms.
authors.affiliation: Mediterranean Agronomic Institute of Chania, P.O. Box 85, Chania 73100, Greece.
authors.forename: Codruta
authors.initials: C
authors.lastname: Ignea
authors.affiliation: Institute of Applied Biosciences - Centre for Research and Technology Hellas (INAB-CERTH), P.O. Box 60361, Thermi, 57001 Thessaloniki, Greece.
authors.forename: Fotini A
authors.initials: FA
authors.lastname: Trikka
authors.affiliation: Institute of Applied Biosciences - Centre for Research and Technology Hellas (INAB-CERTH), P.O. Box 60361, Thermi, 57001 Thessaloniki, Greece.
authors.forename: Alexandros K
authors.initials: AK
authors.lastname: Nikolaidis
authors.affiliation: Department of Pharmacognosy and Chemistry of Natural Products, School of Pharmacy, University of Athens, Panepistimiopolis Zografou, Athens 15771, Greece.
authors.forename: Panagiota
authors.initials: P
authors.lastname: Georgantea
authors.affiliation: Department of Pharmacognosy and Chemistry of Natural Products, School of Pharmacy, University of Athens, Panepistimiopolis Zografou, Athens 15771, Greece.
authors.forename: Efstathia
authors.initials: E
authors.lastname: Ioannou
authors.affiliation: Mediterranean Agronomic Institute of Chania, P.O. Box 85, Chania 73100, Greece.
authors.forename: Sofia
authors.initials: S
authors.lastname: Loupassaki
authors.affiliation: Mediterranean Agronomic Institute of Chania, P.O. Box 85, Chania 73100, Greece.
authors.forename: Panagiotis
authors.initials: P
authors.lastname: Kefalas
authors.affiliation: Department of Pharmaceutical Sciences, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece.
authors.forename: Angelos K
authors.initials: AK
authors.lastname: Kanellis
authors.affiliation: Department of Pharmacognosy and Chemistry of Natural Products, School of Pharmacy, University of Athens, Panepistimiopolis Zografou, Athens 15771, Greece.
authors.forename: Vassilios
authors.initials: V
authors.lastname: Roussis
authors.affiliation: Institute of Applied Biosciences - Centre for Research and Technology Hellas (INAB-CERTH), P.O. Box 60361, Thermi, 57001 Thessaloniki, Greece.
authors.forename: Antonios M
authors.initials: AM
authors.lastname: Makris
authors.affiliation: Mediterranean Agronomic Institute of Chania, P.O. Box 85, Chania 73100, Greece; Department of Biochemistry, School of Medicine, University of Crete, P.O. Box 2208, Heraklion 71003, Greece. Electronic address: s.kampranis@med.uoc.gr.
authors.forename: Sotirios C
authors.initials: SC
authors.lastname: Kampranis
mesh_terms: D004224:Diterpenes / Q000378:metabolism*; D051228:Geranyltranstransferase / Q000096:biosynthesis* / Q000235:genetics; D060847:Metabolic Engineering; D011106:Polyisoprenyl Phosphates / Q000096:biosynthesis; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D004224:Diterpenes; D011106:Polyisoprenyl Phosphates; D029701:Saccharomyces cerevisiae Proteins; D051228:Geranyltranstransferase; C002963:geranylgeranyl pyrophosphate
keywords: Geranylgeranyl diphosphate; Isoprenoid; Protein engineering; Terpene synthase; Yeast