| The availability of recombinant proteins repertoires allows to study large numbers of proteins in parallel, e.g. by screening of protein arrays. Parallel approaches for protein expression also allow screening for protein variants with optimal stability and production yield, thereby enabling the structural and functional analysis of challenging proteins. Structural Genomics studies of bacterial proteins have successfully produced large numbers of bacterial proteins in E. coli, followed by production and crystallisation. The same approach is less successful for mammalian proteins, since only a few protein families can be produced as full length, functional proteins in E. coli. For the expression of mammalian proteins in E. coli, a combination of systematic screening and rational design based on specific knowledge on the respective protein family is useful. Mammalian cell lines are excellent for the production of secreted proteins including glycoproteins. Transient transfection of HEK293 cells can be performed in high throughput and can be scaled up to the demands of crystallography projects. Stable cell lines, once established, can be used indefinitely to produce a protein of interest with little technical effort. Site-specific recombination allows to establish highly productive stable CHO cell lines in a matter of weeks. |
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