title: Metabolic engineering of β-oxidation in Penicillium chrysogenum for improved semi-synthetic cephalosporin biosynthesis.
abstract: Industrial production of semi-synthetic cephalosporins by Penicillium chrysogenum requires supplementation of the growth media with the side-chain precursor adipic acid. In glucose-limited chemostat cultures of P. chrysogenum, up to 88% of the consumed adipic acid was not recovered in cephalosporin-related products, but used as an additional carbon and energy source for growth. This low efficiency of side-chain precursor incorporation provides an economic incentive for studying and engineering the metabolism of adipic acid in P. chrysogenum. Chemostat-based transcriptome analysis in the presence and absence of adipic acid confirmed that adipic acid metabolism in this fungus occurs via β-oxidation. A set of 52 adipate-responsive genes included six putative genes for acyl-CoA oxidases and dehydrogenases, enzymes responsible for the first step of β-oxidation. Subcellular localization of the differentially expressed acyl-CoA oxidases and dehydrogenases revealed that the oxidases were exclusively targeted to peroxisomes, while the dehydrogenases were found either in peroxisomes or in mitochondria. Deletion of the genes encoding the peroxisomal acyl-CoA oxidase Pc20g01800 and the mitochondrial acyl-CoA dehydrogenase Pc20g07920 resulted in a 1.6- and 3.7-fold increase in the production of the semi-synthetic cephalosporin intermediate adipoyl-6-APA, respectively. The deletion strains also showed reduced adipate consumption compared to the reference strain, indicating that engineering of the first step of β-oxidation successfully redirected a larger fraction of adipic acid towards cephalosporin biosynthesis.
authors.affiliation: Industrial Microbiology Section, Department of Biotechnology, Delft University of Technology, Julianalaan 67, 2628 BC Delft, The Netherlands.
authors.forename: Tânia
authors.initials: T
authors.lastname: Veiga
authors.forename: Andreas K
authors.initials: AK
authors.lastname: Gombert
authors.forename: Nils
authors.initials: N
authors.lastname: Landes
authors.forename: Maarten D
authors.initials: MD
authors.lastname: Verhoeven
authors.forename: Jan A K W
authors.initials: JA
authors.lastname: Kiel
authors.forename: Arjen M
authors.initials: AM
authors.lastname: Krikken
authors.forename: Jeroen G
authors.initials: JG
authors.lastname: Nijland
authors.forename: Hesselien
authors.initials: H
authors.lastname: Touw
authors.forename: Marijke A H
authors.initials: MA
authors.lastname: Luttik
authors.forename: John C
authors.initials: JC
authors.lastname: van der Toorn
authors.forename: Arnold J M
authors.initials: AJ
authors.lastname: Driessen
authors.forename: Roel A L
authors.initials: RA
authors.lastname: Bovenberg
authors.forename: Marco A
authors.initials: MA
authors.lastname: van den Berg
authors.forename: Ida J
authors.initials: IJ
authors.lastname: van der Klei
authors.forename: Jack T
authors.initials: JT
authors.lastname: Pronk
authors.forename: Jean-Marc
authors.initials: JM
authors.lastname: Daran
mesh_terms: D044944:Acyl-CoA Dehydrogenases / Q000235:genetics / Q000378:metabolism; D042962:Acyl-CoA Oxidase / Q000235:genetics / Q000378:metabolism; D000272:Adipates / Q000378:metabolism; D002511:Cephalosporins / Q000096:biosynthesis*; D017353:Gene Deletion; D015966:Gene Expression Regulation, Fungal; D005947:Glucose / Q000378:metabolism; D060847:Metabolic Engineering / Q000379:methods*; D008928:Mitochondria / Q000201:enzymology / Q000235:genetics; D010084:Oxidation-Reduction; D010408:Penicillium chrysogenum / Q000378:metabolism*; D020675:Peroxisomes / Q000201:enzymology / Q000235:genetics; D059467:Transcriptome
chemical_list: D000272:Adipates; D002511:Cephalosporins; C029900:adipic acid; D044944:Acyl-CoA Dehydrogenases; D042962:Acyl-CoA Oxidase; D005947:Glucose