Fountoulakis M
F. Hoffmann-La Roche, Ltd, Pharmaceutical Research-Gene Technologies, Basel, Switzerland.
J Chem Technol Biotechnol. 1996 Feb;65(2):123-30. doi: 10.1002/(SICI)1097-4660(199602)65:2<123::AID-JCTB400>3.0.CO;2-6.
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.
重组蛋白表现出几种类型的异质性和翻译后修饰，这通常与其生产系统有关。已对在大肠杆菌、杆状病毒感染的昆虫细胞和中国仓鼠卵巢细胞中表达的重组干扰素γ受体和干扰素γ受体-免疫球蛋白G融合蛋白的明显异质性进行了研究。一般来说，所有测试的蛋白质都表现出某种类型的异质性，可通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳检测到。大肠杆菌来源的受体包括涉及错配或未形成二硫键的非天然构象。这种类型的异质性影响了蛋白质的生物活性。此外，原核蛋白在下游加工过程中截留了磷酸。磷酸截留不影响配体结合能力。真核蛋白由于信号肽切割不均以及糖基化差异而表现出异质性。后一种类型的异质性不影响活性。糖基化相关的异质性部分源于潜在N-糖基化位点的利用不均，并不同程度地影响了蛋白质在聚丙烯酰胺凝胶上的表观分子量和迁移率。这些结果可能有助于重组蛋白的表征研究。
