title: Molecular and catalytic properties of fungal extracellular cellobiose dehydrogenase produced in prokaryotic and eukaryotic expression systems.
abstract: BACKGROUND
  Cellobiose dehydrogenase (CDH) is an extracellular enzyme produced by lignocellulolytic fungi. cdh gene expression is high in cellulose containing media, but relatively low CDH concentrations are found in the supernatant of fungal cultures due to strong binding to cellulose. Therefore, heterologous expression of CDH in Pichia pastoris was employed in the last 15 years, but the obtained enzymes were over glycosylated and had a reduced specific activity.
  
  
  RESULTS
  We compare the well-established CDH expression host P. pastoris with the less frequently used hosts Escherichia coli, Aspergillus niger, and Trichoderma reesei. The study evaluates the produced quantity and protein homogeneity of Corynascus thermophilus CDH in the culture supernatants, the purification, and finally compares the enzymes in regard to cofactor loading, glycosylation, catalytic constants and thermostability.
  
  
  CONCLUSIONS
  Whereas E. coli could only express the catalytic dehydrogenase domain of CDH, all eukaryotic hosts could express full length CDH including the cytochrome domain. The CDH produced by T. reesei was most similar to the CDH originally isolated from the fungus C. thermophilus in regard to glycosylation, cofactor loading and catalytic constants. Under the tested experimental conditions the fungal expression hosts produce CDH of superior quality and uniformity compared to P. pastoris.
authors.affiliation: Department of Food Sciences and Technology, Vienna Institute of Biotechnology, BOKU-University of Natural Resources and Life Sciences, Vienna, Austria.
authors.forename: Su
authors.initials: S
authors.lastname: Ma
authors.affiliation: Department of Food Sciences and Technology, Vienna Institute of Biotechnology, BOKU-University of Natural Resources and Life Sciences, Vienna, Austria.
authors.forename: Marita
authors.initials: M
authors.lastname: Preims
authors.affiliation: UMR BDR, INRA, ENVA, Université Paris Saclay, 78350, Jouy en Josas, France.
authors.forename: François
authors.initials: F
authors.lastname: Piumi
authors.affiliation: Research Area Biochemical Technology, Institute of Chemical Engineering, TU Wien, Gumpendorferstrasse 1a, Vienna, Austria.
authors.forename: Lisa
authors.initials: L
authors.lastname: Kappel
authors.affiliation: Research Area Biochemical Technology, Institute of Chemical Engineering, TU Wien, Gumpendorferstrasse 1a, Vienna, Austria.
authors.forename: Bernhard
authors.initials: B
authors.lastname: Seiboth
authors.affiliation: Aix Marseille Université, INRA, BBF (Biodiversité et Biotechnologie Fongiques), Marseille, France.
authors.forename: Eric
authors.initials: E
authors.lastname: Record
authors.affiliation: Department of Food Sciences and Technology, Vienna Institute of Biotechnology, BOKU-University of Natural Resources and Life Sciences, Vienna, Austria.
authors.forename: Daniel
authors.initials: D
authors.lastname: Kracher
authors.affiliation: Department of Food Sciences and Technology, Vienna Institute of Biotechnology, BOKU-University of Natural Resources and Life Sciences, Vienna, Austria. roland.ludwig@boku.ac.at.
authors.forename: Roland
authors.identifier: http://orcid.org/0000-0002-5058-5874
authors.initials: R
authors.lastname: Ludwig
mesh_terms: D001234:Aspergillus niger / Q000201:enzymology / Q000235:genetics*; D002237:Carbohydrate Dehydrogenases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D002384:Catalysis; D003470:Culture Media / Q000737:chemistry; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D015870:Gene Expression; D006031:Glycosylation; D007700:Kinetics; D010843:Pichia / Q000201:enzymology / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism; D020037:Sordariales / Q000201:enzymology; D013696:Temperature; D014242:Trichoderma / Q000201:enzymology / Q000235:genetics*
chemical_list: D003470:Culture Media; D011994:Recombinant Proteins; D002237:Carbohydrate Dehydrogenases; C019859:cellobiose-quinone oxidoreductase
keywords: Aspergillus niger; Cellobiose dehydrogenase; Cofactor loading; Escherichia coli; Glycoforms; Heterologous expression; Pichia pastoris; Trichoderma reesei