Skip to main content
Kent Academic Repository

Posttranslational processing of recombinant human interferon-gamma in animal expression systems

James, David C., Goldman, Merlin H., Hoare, Michael, Jenkins, Nigel, Oliver, Ronald W. A., Green, Brian N., Freedman, Robert B. (1996) Posttranslational processing of recombinant human interferon-gamma in animal expression systems. Protein Science, 5 (2). pp. 331-340. ISSN 0961-8368. (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) (KAR id:19260)

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.

Abstract

We have characterized the heterogeneity of recombinant human interferon-gamma (IFN-gamma) produced by three expression systems: Chinese hamster ovary cells, the mammary gland of transgenic mice, and baculovirus-infected Spodoptera frugiperda (Sf9) insect cells. Analyses of whole IFN-gamma proteins by electrospray ionization-mass spectrometry (ESI-MS) from each recombinant source revealed heterogeneous populations of IFN-gamma molecules resulting from variations in N-glycosylation and C-terminal polypeptide cleavages. A series of more specific analyses assisted interpretation of maximum entropy deconvoluted ESI-mass spectra of whole IFN-gamma proteins; MALDI-MS analyses of released, desialylated N-glycans and of deglycosylated IFN-gamma polypeptides were combined with analyses of 2-aminobenzamide labeled sialylated N-glycans by cation-exchange high-performance liquid chromatography. These analyses enabled identification of specific polypeptide cleavage sites and characterization of associated N-glycans. Production of recombinant IFN-gamma in the mammalian expression systems yielded polypeptides C-terminally truncated at dibasic amino acid sites. Mammalian cell derived IFN-gamma molecules displayed oligosaccharides with monosaccharide compositions equivalent to complex, sialylated, or high-mannose type N-glycans. In contrast, IFN-gamma derived from baculovirus-infected Sf9 insect cells was truncated further toward the C-terminus and was associated with neutral (nonsialylated) N-glycans. These data demonstrate the profound influence of host cell type on posttranslational processing of recombinant proteins produced in eukaryotic systems.

Item Type: Article
Subjects: Q Science > QP Physiology (Living systems) > QP517 Biochemistry
Divisions: Divisions > Division of Natural Sciences > Biosciences
Depositing User: R.F. Xu
Date Deposited: 04 Jun 2009 16:10 UTC
Last Modified: 16 Nov 2021 09:57 UTC
Resource URI: https://kar.kent.ac.uk/id/eprint/19260 (The current URI for this page, for reference purposes)

University of Kent Author Information

James, David C..

Creator's ORCID:
CReDIT Contributor Roles:

Freedman, Robert B..

Creator's ORCID:
CReDIT Contributor Roles:
  • Depositors only (login required):

Total unique views for this document in KAR since July 2020. For more details click on the image.