Degradation of an Fc-fusion recombinant protein by host cell proteases: Identification of a CHO cathepsin D protease
Article first published online: 4 AUG 2009
Copyright © 2009 Wiley Periodicals, Inc.
Biotechnology and Bioengineering
Volume 104, Issue 6, pages 1132–1141, 15 December 2009
How to Cite
Robert, F., Bierau, H., Rossi, M., Agugiaro, D., Soranzo, T., Broly, H. and Mitchell-Logean, C. (2009), Degradation of an Fc-fusion recombinant protein by host cell proteases: Identification of a CHO cathepsin D protease. Biotechnol. Bioeng., 104: 1132–1141. doi: 10.1002/bit.22494
- Issue published online: 22 OCT 2009
- Article first published online: 4 AUG 2009
- Accepted manuscript online: 4 AUG 2009 12:00AM EST
- Manuscript Accepted: 27 JUL 2009
- Manuscript Revised: 9 JUL 2009
- Manuscript Received: 29 JAN 2009
- CHO cell culture;
- cathepsin D
A host-cell-related proteolytic activity was identified in a recombinant Fc-fusion protein production process. This report describes the strategy applied to characterize and isolate the enzyme responsible for this degradation by combining cell culture investigation and dedicated analytical tools. After isolation and sequencing of the clipped fragment generated in post-capture material, enzymatic activity was traced in different culture conditions, allowing identification of viable CHO cells as the source of protease. Inhibitors and pH screenings showed that the enzyme belongs to an aspartic protease family and is preferably active at acidic pH. The protease was isolated by purification on a pepstatin A column and characterized as a protein related to cathepsin D. An additional metallo-protease inhibited by EDTA was identified with an optimum activity at neutral pH. This study is an example of how quality and stability of therapeutic recombinant molecules are strongly influenced by cell culture parameters. Biotechnol. Bioeng. 2009; 104: 1132–1141. © 2009 Wiley Periodicals, Inc.