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Cloning, expression and purification of three Chaperonin 60 homologues

Journal of Chromatography B, ISSN: 1570-0232, Vol: 786, Issue: 1, Page: 117-125
2003
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Article Description

The Chaperonin 60 (Cpn60) proteins have, in addition to their well-known functions of protein folding and protection, a range of intercellular signalling activities. As part of a study to investigate the biological activity of the Cpn60 proteins, particularly from pathogenic organisms, we have cloned and expressed three Cpn60 proteins from Homo sapiens, Helicobacter pylori and Chlamydia pneumoniae. The Cpn60 proteins were purified to apparent homogeneity using a combination of nickel column affinity chromatography and Reactive Red dye affinity columns. Insoluble protein was solubilised using 8 M urea and then re-folded on the nickel column by stepwise removal of the urea. The immunostimulant LPS was removed by addition of the antibiotic polymyxin B as part of the purification process.

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