Untitled Document
 
 
P05-066: HETEROLOGOUS EXPRESSION OF HORDEUM VULGARE CYSTEINE PROTEASE IN YEAST
 
Authors: Rosenkilde A, (Aarhus University, Department of Genetics and Biotechnology, Faculty of Agricultural Sciences)
  Giuseppe Dionisio, (Aarhus University, Department of Genetics and Biotechnology, Faculty of Agricultural Sciences (DJF),)
  Preben B. Holm, (Aarhus University, Department of Genetics and Biotechnology, Faculty of Agricultural Sciences (DJF),)
  Henrik Brinch-Pedersen, (Aarhus University, Department of Genetics and Biotechnology, Faculty of Agricultural Sciences (DJF),)
 
Topic: Biotechnology
 
Abstract Text:

Cysteine Proteases accounts for more than 90 % of the total proteolytic activity in the degradation of
barley seed storage proteins during germination. Several Cysteine proteases have been identified in
barley. One of the key enzymes, Hordeum vulgare endoprotease B2 (HvEPB2) was cloned with and
without the 5 amino acid C-terminal sequence into the Pichia pastoris expression vector pPICZ Aα and
electrotransformed into Pichia pastoris strain SDM1163. Heterologous protein production was
induced with 2% MeOH. To monitor the protein expression during induction, 1 ml samples was
collected every hr for 24 hrs. After 4 days, the supernatant were harvested and analyzed by SDSPAGE,
activity assay and Western blot. A significant amount of heterologous protein was produced
and the protein production was highest after 4 days and the expression in the C-terminal mutant was
slightly higher than for the full length protease.

 
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