title: Synthetic biology tools for bioprospecting of natural products in eukaryotes.
abstract: Filamentous fungi have the capacity to produce a battery of natural products of often unknown function, synthesized by complex metabolic pathways. Unfortunately, most of these pathways appear silent, many in intractable organisms, and their products consequently unidentified. One basic challenge is the difficulty of expressing a biosynthesis pathway for a complex natural product in a heterologous eukaryotic host. Here, we provide a proof-of concept solution to this challenge and describe how the entire penicillin biosynthesis pathway can be expressed in a heterologous host. The method takes advantage of a combination of improved yeast in vivo cloning technology, generation of polycistronic mRNA for the gene cluster under study, and an amenable and easily manipulated fungal host, i.e., Aspergillus nidulans. We achieve expression from a single promoter of the pathway genes to yield a large polycistronic mRNA by using viral 2A peptide sequences to direct successful cotranslational cleavage of pathway enzymes.
authors.affiliation: Department of Molecular and Applied Microbiology, Leibniz Institute for Natural Product Research and Infection Biology-Hans Knöll Institute (HKI), Beutenbergstrasse 11a, Jena 07745, Germany; School of Biology, Biomedical Sciences Research Complex, University of St. Andrews, St. Andrews, Fife, KY16 9ST, UK. Electronic address: su@st-andrews.ac.uk.
authors.forename: Shiela E
authors.initials: SE
authors.lastname: Unkles
authors.affiliation: Department of Molecular and Applied Microbiology, Leibniz Institute for Natural Product Research and Infection Biology-Hans Knöll Institute (HKI), Beutenbergstrasse 11a, Jena 07745, Germany.
authors.forename: Vito
authors.initials: V
authors.lastname: Valiante
authors.affiliation: Department of Molecular and Applied Microbiology, Leibniz Institute for Natural Product Research and Infection Biology-Hans Knöll Institute (HKI), Beutenbergstrasse 11a, Jena 07745, Germany; Department of Microbiology and Molecular Biology, Institute of Microbiology, Friedrich Schiller University Jena, Beutenbergstrasse 11a, Jena 07745, Germany.
authors.forename: Derek J
authors.initials: DJ
authors.lastname: Mattern
authors.affiliation: Department of Molecular and Applied Microbiology, Leibniz Institute for Natural Product Research and Infection Biology-Hans Knöll Institute (HKI), Beutenbergstrasse 11a, Jena 07745, Germany; Department of Microbiology and Molecular Biology, Institute of Microbiology, Friedrich Schiller University Jena, Beutenbergstrasse 11a, Jena 07745, Germany. Electronic address: Axel.Brakhage@hki-jena.de.
authors.forename: Axel A
authors.initials: AA
authors.lastname: Brakhage
mesh_terms: D001233:Aspergillus nidulans / Q000166:cytology / Q000235:genetics / Q000378:metabolism; D001688:Biological Products / Q000737:chemistry / Q000378:metabolism*; D056890:Eukaryota / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D005810:Multigene Family / Q000235:genetics; D010406:Penicillins / Q000096:biosynthesis* / Q000737:chemistry; D011401:Promoter Regions, Genetic / Q000235:genetics; D064210:Secondary Metabolism; D058615:Synthetic Biology / Q000379:methods*; D014764:Viral Proteins / Q000378:metabolism
chemical_list: D001688:Biological Products; D010406:Penicillins; D014764:Viral Proteins