Studies to prevent degradation of recombinant fc-fusion protein expressed in mammalian cell line and protein characterization Chakrabarti, Sanjukta, Barrow, Colin J., Kanwar, Rupinder K., Ramana, Venkata and Kanwar, Jagat R. 2016, Studies to prevent degradation of recombinant fc-fusion protein expressed in mammalian cell line and protein characterization, International journal of molecular sciences, vol. 17, no. 6, pp. 1-22, doi: 10.3390/ijms17060913. Attached Files Name Description MIMEType Size Downloads barrow-studiestoprevent-2016.pdf Published version application/pdf 4.01MB 17 Title Studies to prevent degradation of recombinant fc-fusion protein expressed in mammalian cell line and protein characterization Author(s) Chakrabarti, Sanjukta Barrow, Colin J. orcid.org/0000-0002-2153-7267 Kanwar, Rupinder K. Ramana, Venkata Kanwar, Jagat R. orcid.org/0000-0003-3728-9568 Journal name International journal of molecular sciences Volume number 17 Issue number 6 Article ID 913 Start page 1 End page 22 Total pages 22 Publisher MDPI Place of publication Basel, Switzerland. Publication date 2016-06 ISSN 1422-0067 Keyword(s) CHOK1SV GS-KO Fc fusion protein VEGFR1(D1–D3)-Fc biological activity clipping folded inhibition proteolysis stable Science & Technology Life Sciences & Biomedicine Physical Sciences Biochemistry & Molecular Biology Chemistry, Multidisciplinary Chemistry VEGFR1(D1-D3)-Fc CIRCULAR-DICHROISM SPECTROSCOPY HAMSTER OVARY CELLS HETEROGENEITY PRODUCTIVITY ANTIBODIES PEPTIDES Summary Clipping of recombinant proteins is a major issue in animal cell cultures. A recombinant Fc-fusion protein, VEGFR1(D1-D3)-Fc expressed in CHOK1SV GS-KO cells was observed to be undergoing clippings in lab scale cultures. Partial cleaving of expressed protein initiated early on in cell culture and was observed to increase over time in culture and also on storage. In this study, a few parameters were explored in a bid to inhibit clipping in the fusion protein The effects of culture temperature, duration of culture, the addition of an anti-clumping agent, ferric citrate and use of protease inhibitor cocktail on inhibition of proteolysis of the Fc fusion were studied. Lowering of culture temperature from 37 to 30 °C alone appears to be the best solution for reducing protein degradation from the quality, cost and regulatory points of view. The obtained Fc protein was characterized and found to be in its stable folded state, exhibiting a high affinity for its ligand and also biological and functional activities. Language eng DOI 10.3390/ijms17060913 Field of Research 119999 Medical and Health Sciences not elsewhere classified 0399 Other Chemical Sciences 0604 Genetics 0699 Other Biological Sciences Socio Economic Objective 929999 Health not elsewhere classified HERDC Research category C1 Refereed article in a scholarly journal ERA Research output type C Journal article Copyright notice ©2016, The Authors Free to Read? Yes Use Rights Persistent URL http://hdl.handle.net/10536/DRO/DU:30085982 Document type: Journal Article Collections: School of Medicine School of Life and Environmental Sciences Open Access Collection Related Links Link Description Connect to published version Go to link with your DU access privileges     Unless expressly stated otherwise, the copyright for items in DRO is owned by the author, with all rights reserved. Every reasonable effort has been made to ensure that permission has been obtained for items included in DRO. If you believe that your rights have been infringed by this repository, please contact drosupport@deakin.edu.au. Versions Version Filter Type Wed, 07 Sep 2016, 14:24:57 EST Thu, 08 Sep 2016, 05:01:27 EST Thu, 08 Sep 2016, 06:11:35 EST Mon, 07 Nov 2016, 12:40:18 EST Mon, 07 Nov 2016, 12:43:37 EST Tue, 08 Nov 2016, 14:52:21 EST Wed, 09 Nov 2016, 12:38:56 EST Wed, 09 Nov 2016, 12:42:13 EST Wed, 09 Nov 2016, 12:46:37 EST Tue, 18 Jul 2017, 11:16:56 EST Fri, 07 Sep 2018, 12:40:18 EST Filtered Full Citation counts:   Cited 3 times in TR Web of Science Cited 3 times in Scopus Search Google Scholar Access Statistics: 226 Abstract Views, 17 File Downloads  -  Detailed Statistics Created: Wed, 07 Sep 2016, 14:24:57 EST

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