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J Struct Biol. 2010 Oct;172(1):3-13. doi: 10.1016/j.jsb.2010.06.008. Epub 2010 Jun 10.

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

Author information

1
Structural Genomics Consortium, University of Oxford, Old Road Campus Research Building, Oxford, UK. pavel.savitsky@sgc.ox.ac.uk

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 approximately 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.

PMID:
20541610
PMCID:
PMC2938586
DOI:
10.1016/j.jsb.2010.06.008
[Indexed for MEDLINE]
Free PMC Article

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