Expression of lignocellulolytic enzymes in Pichia pastoris

Andrea Mellitzer, Roland Weis, Anton Glieder, Karlheinz Flicker

Research output: Contribution to journalArticle

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.

CONCLUSION: 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.

Original languageEnglish
Pages (from-to)61
Number of pages11
JournalMicrobial cell factories
Volume11
DOIs
Publication statusPublished - 14 May 2012

Keywords

  • Alcohol Oxidoreductases
  • Cellulose 1,4-beta-Cellobiosidase
  • Endo-1,4-beta Xylanases
  • Fermentation
  • Gene Dosage
  • Gene Expression Regulation, Fungal
  • Pichia
  • Promoter Regions, Genetic
  • Recombinant Proteins
  • Trichoderma
  • beta-Mannosidase
  • Journal Article
  • Research Support, Non-U.S. Gov't

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