Apparent heterogeneity of recombinant interferon gamma receptors produced in prokaryotic and eukaryotic expression systems.
Recombinant proteins show several types of heterogeneity and post-translational modifications which are usually related to their production system. The apparent heterogeneity of recombinant interferon gamma receptors and interferon gamma receptor-immunoglobulin G fusion proteins expressed in Escherichia coli, baculovirus-infected insect cells and Chinese hamster ovary cells have been studied. In general, all proteins tested showed some type of heterogeneity which was detectable by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The E. coli-derived receptor included non-native conformations involving mis-paired or non-formed disulfides. This type of heterogeneity affected the biological activity of the protein. In addition, the prokaryotic protein had trapped phosphoric acid during downstream processing. The phosphoric acid entrapment did not affect ligand binding capacity. The eukaryotic proteins showed heterogeneity because of the unequal cleavage of the signal peptide and because of differences in glycosylation. The latter types of heterogeneity did not affect activity. Glycosylation-related heterogeneity was partially derived from the unequal utilization of the potential N-glycosylation sites and differently affected the apparent molecular masses and migrations of the proteins on polyacrylamide gels. The results may be useful in characterization studies of recombinant proteins.
Fountoulakis
M
M
F. Hoffmann-La Roche, Ltd, Pharmaceutical Research-Gene Technologies, Basel, Switzerland.
D000595:Amino Acid Sequence; D000818:Animals; D015703:Antigens, CD / Q000096:biosynthesis* / Q000737:chemistry* / Q000302:isolation & purification; D001709:Biotechnology; D016466:CHO Cells; D003001:Cloning, Molecular / Q000379:methods; D006224:Cricetinae; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli; D006031:Glycosylation; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D017471:Receptors, Interferon / Q000096:biosynthesis* / Q000737:chemistry* / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification; D014162:Transfection; D000096743:Interferon gamma Receptor
D015703:Antigens, CD; D017471:Receptors, Interferon; D011994:Recombinant Proteins; D000096743:Interferon gamma Receptor