Evaluation of different expression systems for the heterologous expression of pyranose 2-oxidase from Trametes multicolor in E. coli.
The heterologous production of the industrially relevant fungal enzyme pyranose 2-oxidase in the prokaryotic host E. coli was investigated using 3 different expression systems, i.e. the well-studied T7 RNA polymerase based pET21d+, the L-arabinose inducible pBAD and the pCOLD system. Preliminary experiments were done in shaking flasks at 25 degrees C and optimized induction conditions to compare the productivity levels of the different expression systems. The pET21d+ and the pCOLD system gave 29 U/L x h and 14 U/L x h of active pyranose 2-oxidase, respectively, whereas the pBAD system only produced 6 U/L x h. Process conditions for batch fermentations were optimized for the pET21d+ and the pCOLD systems in order to reduce the formation of inactive inclusion bodies. The highest productivity rate with the pET21d+ expression system in batch fermentations was determined at 25 degrees C with 32 U/L x h. The pCOLD system showed the highest productivity rate (19 U/L x h) at 25 degrees C and induction from the start of the cultivation. Using the pCOLD system in a fed batch fermentation at 25 degrees C with a specific growth rate of mu = 0.15 h-1resulted in the highest productivity rate of active pyranose oxidase with 206 U/L x h.
Spadiut
Oliver
O
Food Biotechnology Lab, Department of Food Sciences and Technology, BOKU - University of Natural Resources and Applied Life Sciences Vienna, Austria.
Posch
Gerald
G
Ludwig
Roland
R
Haltrich
Dietmar
D
Peterbauer
Clemens K
CK
D002237:Carbohydrate Dehydrogenases / Q000235:genetics* / Q000378:metabolism; D046508:Culture Techniques; D004926:Escherichia coli / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D005818:Genetic Engineering / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D055454:Trametes / Q000201:enzymology*
D005656:Fungal Proteins; D002237:Carbohydrate Dehydrogenases; C022489:pyranose oxidase