High-throughput production of human proteins for crystallization: The SGC experience

Pavel Savitsky, James Bray, Christopher D O Cooper, Brian D. Marsden, Pravin Mahajan, Nicola A. Burgess-Brown, Opher Gileadi

Research output: Contribution to journalArticle

176 Citations (Scopus)

Abstract

Producing purified human proteins with high yield and purity remains a considerable challenge. We describe the methods utilized in the Structural Genomics Consortium (SGC) in Oxford, resulting in successful purification of 48% of human proteins attempted; of those, the structures of ∼40% were solved by X-ray crystallography. The main driver has been the parallel processing of multiple (typically 9-20) truncated constructs of each target; modest diversity in vectors and host systems; and standardized purification procedures. We provide method details as well as data on the properties of the constructs leading to crystallized proteins and the impact of methodological variants. These can be used to formulate guidelines for initial approaches to expression of new eukaryotic proteins.

Original languageEnglish
Pages (from-to)3-13
Number of pages11
JournalJournal of Structural Biology
Volume172
Issue number1
DOIs
Publication statusPublished - Oct 2010
Externally publishedYes

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    Savitsky, P., Bray, J., Cooper, C. D. O., Marsden, B. D., Mahajan, P., Burgess-Brown, N. A., & Gileadi, O. (2010). High-throughput production of human proteins for crystallization: The SGC experience. Journal of Structural Biology, 172(1), 3-13. https://doi.org/10.1016/j.jsb.2010.06.008