Recovery and reuse of the molecular chaperone GroEL for in vitro protein refolding

Biotechnology Progress
A D Guise, J B Chaudhuri

Abstract

The chaperones GroEL and GroES from Escherichia coli are known to improve in vitro protein refolding yields. We show that, for the molecular chaperone-assisted refolding of hen egg white lysozyme, GroES is not an essential requirement and that activity is recovered with GroEL and ATP alone. The refolding yields of lysozyme in the presence of GroEL are much greater than those obtained by dilution because of a reduction in protein aggregation. On the basis of the large difference in molecular weight between the GroEL complex (MW 840 000) and lysozyme (MW 14 600), we have demonstrated that using an ultrafiltration membrane (MW 30 000) GroEL may be easily retained after refolding while lysozyme passes freely into the permeate. The chaperonin recovered from the refolding solution was then reused several times for further refolding experiments. The effectiveness of GroEL-assisted refolding was found to decrease with reuse, and this has been attributed to a reduction in the GroEL:lysozyme molar ratio.

Citations

Jun 24, 2000·Journal of Chromatography. a·X Y DongY Sun
Nov 19, 2009·Chemical Communications : Chem Comm·Jun OkadaMasahiro Goto
Oct 20, 2005·Journal of Bioscience and Bioengineering·T MannenT Nagamune
Oct 20, 2005·Journal of Bioscience and Bioengineering·Masafumi SakonoMasahiro Goto
Oct 13, 2006·Acta Biochimica Et Biophysica Sinica·Zhen-Hua YongDa-Quan Xu

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