Immobilisation of bovine enterokinase and application of the immobilised enzyme in fusion protein cleavage

Bioprocess and Biosystems Engineering
Tina KubitzkiStephan Lütz

Abstract

Two immobilisation methods for enterokinase were developed, which yielded high remaining activities for the cleavage of the fusion protein MUC1-IgG Fc. Different carrier materials were compared regarding remaining enzyme activity and storage stability. Immobilisation procedures involving support material activation using glutardialdehyde were found to result in low remaining activities. Applying less aggressive activation procedures, remaining activities of approximately 60% were received when immobilising enterokinase on either Estapor paramagnetic microspheres or hexamethylamino Sepabeads. In case of hexamethylamino Sepabeads we were able to increase the half-life time 4.3-fold at 23 degrees C and 3.8-fold at 4 degrees C compared to the free enzyme at the same temperatures. By immobilising the biocatalyst the downstream process is simplified allowing the easy removal of the enzyme from the reaction mixture. The immobilised enterokinase cleaves the fusion protein MUC1-IgG Fc in at least two repeated batches, proving the efficiency of the immobilisation method and the reusability of the biocatalyst.

References

May 15, 1972·Comparative Biochemistry and Physiology. B, Comparative Biochemistry·S Bricteux-GrégoireM Florkin
Feb 14, 1980·Clinica Chimica Acta; International Journal of Clinical Chemistry·I AntonowiczB Hadorn
Mar 1, 1996·International Journal of Cancer. Journal International Du Cancer·R A GrahamJ Taylor-Papadimitriou
Mar 13, 2003·Biotechnology and Applied Biochemistry·Chang Woo SuhEun Kyu Lee
Mar 27, 2004·Life Sciences·Junling LiKeith B Glaser
Aug 27, 2004·Biotechnology and Bioengineering·R SchoevaartR A Sheldon
Nov 20, 2004·Annals of Oncology : Official Journal of the European Society for Medical Oncology·J S de BonoA W Tolcher
Jan 1, 1982·Biophysical Journal·P Fonseca, A Light
Mar 20, 1939·The Journal of General Physiology·M Kunitz
Mar 20, 1939·The Journal of General Physiology·M Kunitz

❮ Previous
Next ❯

Citations

Mar 21, 2008·Bioprocess and Biosystems Engineering·Dirk Weuster-Botz
Dec 7, 2011·The FEBS Journal·Patrick R StancombeK Ravi Acharya
Jul 24, 2012·Journal of Biotechnology·Sara D F SantanaAna C A Roque
Feb 4, 2012·Journal of Biotechnology·Baligh MiladiAbdellatif Elm'selmi
Sep 3, 2011·Journal of Biotechnology·Stanislav PepeliaevJana Dostálová
Aug 30, 2011·International Journal of Biological Macromolecules·Hasan BayraktarSeçil Onal

❮ Previous
Next ❯

Related Concepts

Related Feeds

Bioinformatics in Biomedicine

Bioinformatics in biomedicine incorporates computer science, biology, chemistry, medicine, mathematics and statistics. Discover the latest research on bioinformatics in biomedicine here.