title: Constitutive homologous expression of phosphoglucomutase and transaldolase increases the metabolic flux of Fusarium oxysporum.
abstract: BACKGROUND
  Fusarium oxysporum is among the few filamentous fungi that have been reported of being able to directly ferment biomass to ethanol in a consolidated bioprocess. Understanding its metabolic pathways and their limitations can provide some insights on the genetic modifications required to enhance its growth and subsequent fermentation capability. In this study, we investigated the hypothesis reported previously that phosphoglucomutase and transaldolase are metabolic bottlenecks in the glycolysis and pentose phosphate pathway of the F. oxysporum metabolism.
  
  
  RESULTS
  Both enzymes were homologously overexpressed in F. oxysporum F3 using the gpdA promoter of Aspergillus nidulans for constitutive expression. Transformants were screened for their phosphoglucomutase and transaldolase genes expression levels with northern blot. The selected transformant exhibited high mRNA levels for both genes, as well as higher specific activities of the corresponding enzymes, compared to the wild type. It also displayed more than 20 and 15% higher specific growth rate upon aerobic growth on glucose and xylose, respectively, as carbon sources and 30% higher biomass to xylose yield. The determination of the relative intracellular amino and non-amino organic acid concentrations at the end of growth on glucose revealed higher abundance of most determined metabolites between 1.5- and 3-times in the recombinant strain compared to the wild type. Lower abundance of the determined metabolites of the Krebs cycle and an 68-fold more glutamate were observed at the end of the cultivation, when xylose was used as carbon source.
  
  
  CONCLUSIONS
  Homologous overexpression of phosphoglucomutase and transaldolase in F. oxysporum was shown to enhance the growth characteristics of the strain in both xylose and glucose in aerobic conditions. The intracellular metabolites profile indicated how the changes in the metabolome could have resulted in the observed growth characteristics.
authors.forename: George E
authors.initials: GE
authors.lastname: Anasontzis
authors.forename: Elisavet
authors.initials: E
authors.lastname: Kourtoglou
authors.forename: Diomi
authors.initials: D
authors.lastname: Mamma
authors.forename: Silas G
authors.initials: SG
authors.lastname: Villas-Boâs
authors.forename: Dimitris G
authors.initials: DG
authors.lastname: Hatzinikolaou
authors.affiliation: Biochemical and Chemical Process Engineering, Division of Sustainable Process Engineering, Department of Civil, Environmental and Natural Resources Engineering, Luleå University of Technology, Luleå SE-97187, Sweden. paul.christakopoulos@ltu.se.
authors.forename: Paul
authors.initials: P
authors.lastname: Christakopoulos
mesh_terms: D001233:Aspergillus nidulans / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics; D018533:Biomass; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D005670:Fusarium / Q000254:growth & development / Q000378:metabolism*; D005947:Glucose / Q000378:metabolism; D060847:Metabolic Engineering; D010733:Phosphoglucomutase / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D014153:Transaldolase / Q000235:genetics / Q000378:metabolism*; D014994:Xylose / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D005656:Fungal Proteins; D012333:RNA, Messenger; D014994:Xylose; D014153:Transaldolase; D010733:Phosphoglucomutase; D005947:Glucose