Expressing the human proteome for affinity proteomics: Optimising expression of soluble protein domains and in vivo biotinylation

Tracy Keates, Christopher D O Cooper, Pavel Savitsky, Charles K. Allerston, Claire Phillips, Martin Hammarström, Neha Daga, Georgina Berridge, Pravin Mahajan, Nicola A. Burgess-Brown, Susanne Müller, Susanne Gräslund, Opher Gileadi

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22 Citations (Scopus)

Abstract

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.

Original languageEnglish
Pages (from-to)515-525
Number of pages11
JournalNew Biotechnology
Volume29
Issue number5
DOIs
Publication statusPublished - 15 Jun 2012
Externally publishedYes

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    Keates, T., Cooper, C. D. O., Savitsky, P., Allerston, C. K., Phillips, C., Hammarström, M., Daga, N., Berridge, G., Mahajan, P., Burgess-Brown, N. A., Müller, S., Gräslund, S., & Gileadi, O. (2012). Expressing the human proteome for affinity proteomics: Optimising expression of soluble protein domains and in vivo biotinylation. New Biotechnology, 29(5), 515-525. https://doi.org/10.1016/j.nbt.2011.10.007