Keates, Tracy, Cooper, Christopher D.O., Savitsky, Pavel, Allerston, Charles K., Phillips, Claire, Hammarström, Martin, Daga, Neha, Berridge, Georgina, Mahajan, Pravin, Burgess-Brown, Nicola A., Müller, Susanne, Gräslund, Susanne and Gileadi, Opher (2012) Expressing the human proteome for affinity proteomics: optimising expression of soluble protein domains and in vivo biotinylation. New Biotechnology, 29 (5). pp. 515-525. ISSN 1876-4347

The generation of affinity reagents to large numbers of human proteins depends on the ability to express the target proteins as high-quality antigens. The Structural Genomics Consortium (SGC) focuses on the production and structure determination of human proteins. In a 7-year period, the SGC has deposited crystal structures of >800 human protein domains, and has additionally expressed and purified a similar number of protein domains that have not yet been crystallised. The targets include a diversity of protein domains, with an attempt to provide high coverage of protein families. The family approach provides an excellent basis for characterising the selectivity of affinity reagents. We present a summary of the approaches used to generate purified human proteins or protein domains, a test case demonstrating the ability to rapidly generate new proteins, and an optimisation study on the modification of >70 proteins by biotinylation in vivo. These results provide a unique synergy between large-scale structural projects and the recent efforts to produce a wide coverage of affinity reagents to the human proteome.

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