Büssow Konrad
Helmholtz Centre for Infection Research, Structure and Function of Proteins, Inhoffenstr. 7, 38124 Braunschweig, Germany.
Curr Opin Struct Biol. 2015 Jun;32:81-90. doi: 10.1016/j.sbi.2015.03.002. Epub 2015 Mar 23.
The mammalian cell lines HEK293 and CHO have become important expression hosts in structural biology. Generating stable mammalian cell lines remains essential for studying the function and structure of recombinant proteins, despite the emergence of highly efficient transient transfection protocols. Production with stable cell lines can be scaled up easily and high volumetric product yield can be achieved. Protein structure reports of the past two years that used stable cell lines were surveyed for this review. Well-established techniques and novel approaches for generating stable cell lines and stable cell pools are presented, including cell sorting, site-specific recombination, transposons, the Lentivirus system and phage integrases. Host cell line optimization by endoglycosidase overexpression and sequence-specific genome engineering is highlighted.
哺乳动物细胞系HEK293和CHO已成为结构生物学中重要的表达宿主。尽管出现了高效的瞬时转染方案，但生成稳定的哺乳动物细胞系对于研究重组蛋白的功能和结构仍然至关重要。使用稳定细胞系进行生产可以轻松扩大规模，并可实现高体积产物产量。本综述调查了过去两年使用稳定细胞系的蛋白质结构报告。介绍了用于生成稳定细胞系和稳定细胞池的成熟技术和新方法，包括细胞分选、位点特异性重组、转座子、慢病毒系统和噬菌体整合酶。强调了通过内切糖苷酶过表达和序列特异性基因组工程对宿主细胞系进行优化。
