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Posttranslational processing of recombinant human interferon-gamma in animal expression systems
D C James1, M H Goldman, M Hoare
1Research School of Biosciences, University of Kent, Centerbury, United Kingdom. d.c.james@ukc.ac.uk
Protein Science : a Publication of the Protein Society
|February 1, 1996
Summary
Recombinant interferon-gamma (IFN-gamma) from different expression systems shows variations in N-glycosylation and C-terminal cleavage. Host cell type significantly impacts posttranslational processing of these therapeutic proteins.
Area of Science:
- Biochemistry
- Molecular Biology
- Protein Engineering
Background:
- Recombinant human interferon-gamma (IFN-gamma) is a therapeutic protein.
- Understanding posttranslational modifications is crucial for recombinant protein production.
Purpose of the Study:
- To characterize the heterogeneity of recombinant IFN-gamma from three distinct expression systems.
- To investigate the impact of host cell type on N-glycosylation and C-terminal processing.
Main Methods:
- Electrospray ionization-mass spectrometry (ESI-MS) for whole protein analysis.
- MALDI-MS for N-glycan and polypeptide analysis.
- Cation-exchange HPLC for sialylated N-glycan characterization.
Main Results:
- Recombinant IFN-gamma exhibited heterogeneity due to N-glycosylation and C-terminal cleavage variations.
- Mammalian systems produced C-terminally truncated IFN-gamma with complex, sialylated, or high-mannose N-glycans.
- Baculovirus-infected insect cells yielded further truncated IFN-gamma with neutral N-glycans.
Conclusions:
- Host cell type profoundly influences posttranslational processing of recombinant proteins.
- Expression system choice impacts IFN-gamma heterogeneity, affecting its properties.
- These findings are critical for optimizing recombinant protein production for therapeutic use.