title: Expression of lignocellulolytic enzymes in Pichia pastoris.
abstract: BACKGROUND
  Sustainable utilization of plant biomass as renewable source for fuels and chemical building blocks requires a complex mixture of diverse enzymes, including hydrolases which comprise the largest class of lignocellulolytic enzymes. These enzymes need to be available in large amounts at a low price to allow sustainable and economic biotechnological processes.Over the past years Pichia pastoris has become an attractive host for the cost-efficient production and engineering of heterologous (eukaryotic) proteins due to several advantages.
  
  
  RESULTS
  In this paper codon optimized genes and synthetic alcohol oxidase 1 promoter variants were used to generate Pichia pastoris strains which individually expressed cellobiohydrolase 1, cellobiohydrolase 2 and beta-mannanase from Trichoderma reesei and xylanase A from Thermomyces lanuginosus. For three of these enzymes we could develop strains capable of secreting gram quantities of enzyme per liter in fed-batch cultivations. Additionally, we compared our achieved yields of secreted enzymes and the corresponding activities to literature data.
  
  
  CONCLUSIONS
  In our experiments we could clearly show the importance of gene optimization and strain characterization for successfully improving secretion levels. We also present a basic guideline how to correctly interpret the interplay of promoter strength and gene dosage for a successful improvement of the secretory production of lignocellulolytic enzymes in Pichia pastoris.
authors.affiliation: ACIB GmbH, Austrian Centre of Industrial Biotechnology, Graz, Austria.
authors.forename: Andrea
authors.initials: A
authors.lastname: Mellitzer
authors.forename: Roland
authors.initials: R
authors.lastname: Weis
authors.forename: Anton
authors.initials: A
authors.lastname: Glieder
authors.forename: Karlheinz
authors.initials: K
authors.lastname: Flicker
mesh_terms: D000429:Alcohol Oxidoreductases / Q000235:genetics; D043366:Cellulose 1,4-beta-Cellobiosidase / Q000235:genetics / Q000378:metabolism*; D043364:Endo-1,4-beta Xylanases / Q000235:genetics / Q000378:metabolism*; D005285:Fermentation; D018628:Gene Dosage; D015966:Gene Expression Regulation, Fungal; D010843:Pichia / Q000201:enzymology* / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D014242:Trichoderma / Q000201:enzymology; D044902:beta-Mannosidase / Q000235:genetics / Q000378:metabolism*
chemical_list: D011994:Recombinant Proteins; D000429:Alcohol Oxidoreductases; C019643:alcohol oxidase; D044902:beta-Mannosidase; D043364:Endo-1,4-beta Xylanases; D043366:Cellulose 1,4-beta-Cellobiosidase