Dialysis strategies for protein refolding: preparative streptavidin production.
We have investigated different dialysis strategies for the refolding of recombinant streptavidin, and present a novel dialysis setup featuring gradual dilution dialysis and continuous protein feeding into a dialysis sack. A denaturing dialysis buffer is exchanged gradually by dilution with refolding buffer and it is demonstrated that the refolding yield can be increased from 45 to 75% by lowering the dilution rate. In addition, continuous feeding of protein to the dialysis sack increases the yield by 5 to 10%. The principle of gradual dilution dialysis is amenable to stringent regulation and we suggest it to be applied for other insoluble protein targets.
Sørensen
Hans Peter
HP
Laboratory of BioDesign, Department of Molecular Biology, Aarhus University, Gustav Wieds Vej 10 C, DK-8000, Aarhus C, Denmark.
Sperling-Petersen
Hans Uffe
HU
Mortensen
Kim Kusk
KK
D000465:Algorithms; D002850:Chromatography, Gel; D003956:Dialysis / Q000295:instrumentation / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D005822:Genetic Vectors / Q000235:genetics; D019791:Guanidine / Q000737:chemistry; D007544:Isopropyl Thiogalactoside / Q000494:pharmacology; D011489:Protein Denaturation; D017510:Protein Folding; D020836:Protein Structure, Quaternary; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification; D019809:Streptavidin / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics
D011994:Recombinant Proteins; D007544:Isopropyl Thiogalactoside; D019809:Streptavidin; D019791:Guanidine