Overcoming Challenges of Hepatitis C Virus Envelope Glycoprotein Production in Mammalian Cells.
Overcoming Challenges of Hepatitis C Virus Envelope Glycoprotein Production in Mammalian Cells.
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克服哺乳动物细胞中丙型肝炎病毒包膜糖蛋白生产的挑战。
DOI：
10.1007/978-1-4939-8976-8_21
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发表时间：
2019
期刊：
影响因子：
--
通讯作者：
Marcotrigiano,Joseph
中科院分区：
文献类型：
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作者：
Yost,SamanthaA;Whidby,Jillian;Khan,AbdulG;Wang,Yuanyuan;Marcotrigiano,Joseph
Posttranslational modifications (PTMs) are often required for proper folding and physiological function of proteins, including the envelope glycoproteins 1 and 2 (E1 and E2) of hepatitis C virus (HCV). Commonly used expression systems such as bacteria, yeast, and baculovirus produce soluble HCV E1 and E2 at very low yields, as the cellular environment and molecular machinery necessary for PTMs may be suboptimal or missing. Here, we describe an expression system for HCV E2 ectodomain (eE2) with 11 N-linked glycans and eight disulfide bonds, which combines lentivirus transduction of mammalian cells and a continuous growth, adherent cell bioreactor. It is environmentally friendly, as well as cost- and time-efficient compared to other methods of recombinant protein expression in mammalian systems with final yields of eE2 approaching 60 mg/L of cell culture supernatant. eE2 produced by this system is amenable to a variety of biophysical studies, including structural determination by X-ray crystallography. Considering the ease of use and flexibility, this method can be applied to express an array of difficult target proteins in a variety of mammalian cell lines.
