Monitoring proteolysis of recombinant human interferon-gamma during batch culture of Chinese hamster ovary cells

Goldman, Merlin H. and James, David C. and Ison, Andrew P. and Bull, Alan T. (1997) Monitoring proteolysis of recombinant human interferon-gamma during batch culture of Chinese hamster ovary cells. Cytotechnology, 23 (1-3). pp. 103-111. ISSN 0920-9069. (The full text of this publication is not currently available from this repository. You may be able to access a copy if URLs are provided)

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Proteolytic cleavage of recombinant human interferon-gamma (IFN-gamma) expressed in Chinese hamster ovary (CHO) cells during batch fermentation has been monitored by mass spectrometric peptide mapping. IFN-gamma was purified from cell-free culture supernatant by immunoaffinity chromatography and cleaved by endoprotease Asp-N. Peptide fragments were resolved by reverse-phase HPLC and identified by a combination of matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) and automated N-terminal peptide sequencing. Using this approach, a peptide was identified as the C-terminal fragment of the IFN-gamma polypeptide. Analysis of this peptide by MS indicated that the recombinant IFN-gamma polypeptide secreted by CHO cells was truncated by at least ten amino acids, initially at Gln(133)-Met(134). No full length (143 amino acids) polypeptide molecules were observed at any stages of the fermentation. Additional proteolytic cleavages at basic amino acids N-terminal of Gln(133) occurred during the later stages of the culture resulting in a heterogeneous IFN-gamma polypeptide population with 'ragged' C-termini.

Item Type: Article
Uncontrolled keywords: batch culture; Chinese hamster ovary; interferon-gamma; mass spectrometry; proteolytic processing; serum-free cultivation
Subjects: Q Science
Q Science > QP Physiology (Living systems) > QP506 Molecular biology
Divisions: Faculties > Science Technology and Medical Studies > School of Biosciences
Depositing User: M.A. Ziai
Date Deposited: 06 Mar 1914 09:06
Last Modified: 23 Apr 2014 10:55
Resource URI: (The current URI for this page, for reference purposes)
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