Causes of Proteolytic Degradation of Secreted Recombinant Proteins Produced in ethylotrophic Yeast Pichia pastoris: Case Study With Recombinant Ovine Interferon-T

Authors

Jayanta Sinha, Department of Chemical Engineering, University of Nebraska-LincolnFollow
Bradley A. Plantz, Department of Chemical Engineering, University of Nebraska-Lincoln
Mehmet Inan, Department of Chemical Engineering, University of Nebraska-LinconlFollow
Michael Meagher, Department of Chemical Engineering, University of Nebraska-LincolnFollow

Date of this Version

December 2004

Comments

Originally published online 3 December 2004 in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/bit.20318 This article can be viewed at the publishers site.

Abstract

It was observed that during fermentative production of recombinant ovine interferon-H (r-oIFN-H ) in Pichia pastoris, a secreted recombinant protein, the protein was degraded increasingly after 48 h of induction and the rate of degradation increased towards the end of fermentation at 72 h, when the fermentation was stopped. Proteases, whose primary source was the vacuoles, was found in in-creasing levels in the cytoplasm and in the fermentation broth after 48 h of induction and reached maximal values when the batch was completed at 72 h. Protease levels at various cell fractions as well as in the culture supernatant were lower when glycerol was used as the carbon source instead of methanol. It can be concluded that methanol me-tabolism along with cell lysis towards the end of fermenta-tion contributes to increased proteolytic activity and even-tual degradation of recombinant protein