Continuous processing of fusion protein expressed as an Escherichia coli inclusion body.
In this study we develop the components of an integrated process for the continuous extraction and purification of a histidine-tagged fusion protein expressed as an inclusion body in Escherichia coli. Lac21 was selected as a model peptide and was expressed as a fusion to ketosteroid isomerase. A purification strategy was developed on a 1-ml batch column before successful scale-up and transfer to a continuous purification system, having a bed volume of 240 ml. Preliminary experiments proved cleavage of the fusion protein. The use of chemical extraction and continuous chromatography gives a flowsheet far superior to the traditional methods for inclusion body processing.
Morreale
Giacomo
G
Department of Chemical Engineering, University of Cambridge, Pembroke Street, CB2 3RA, Cambridge, UK.
Lanckriet
Heikki
H
Miller
Jemma C
JC
Middelberg
Anton P J
AP
D015153:Blotting, Western; D002853:Chromatography, Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D002479:Inclusion Bodies / Q000378:metabolism*; D010957:Plasmids; D011993:Recombinant Fusion Proteins / Q000378:metabolism*
D011993:Recombinant Fusion Proteins