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Eukaryotic expression: developments for structural proteomics

✍ Scribed by Aricescu, A. R. ;Assenberg, R. ;Bill, R. M. ;Busso, D. ;Chang, V. T. ;Davis, S. J. ;Dubrovsky, A. ;Gustafsson, L. ;Hedfalk, K. ;Heinemann, U. ;Jones, I. M. ;Ksiazek, D. ;Lang, C. ;Maskos, K. ;Messerschmidt, A. ;Macieira, S. ;Peleg, Y. ;Perrakis, A. ;Poterszman, A. ;Schneider, G. ;Sixma, T. K. ;Sussman, J. L. ;Sutton, G. ;Tarboureich, N. ;Zeev-Ben-Mordehai, T. ;Jones, E. Yvonne

Publisher
International Union of Crystallography
Year
2006
Tongue
English
Weight
280 KB
Volume
62
Category
Article
ISSN
0907-4449

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✦ Synopsis

The production of sufficient quantities of protein is an essential prelude to a structure determination, but for many viral and human proteins this cannot be achieved using prokaryotic expression systems. Groups in the Structural Proteomics In Europe (SPINE) consortium have developed and implemented high-throughput (HTP) methodologies for cloning, expression screening and protein production in eukaryotic systems. Studies focused on three systems: yeast (Pichia pastoris and Saccharomyces cerevisiae), baculovirus-infected insect cells and transient expression in mammalian cells. Suitable vectors for HTP cloning are described and results from their use in expression screening and protein-production pipelines are reported. Strategies for co-expression, selenomethionine labelling (in all three eukaryotic systems) and control of glycosylation (for secreted proteins in mammalian cells) are assessed.

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