Procedure for refolding and purification of recombinant proteins from Escherichia coli inclusion bodies using a strong anion exchanger.
Using Escherichia coli system expressing papilloma virus HPV16 E7MS2 fusion protein as a model system, a novel procedure was applied to solubilize, purify and refold recombinant proteins from E. coli inclusion bodies. The necessity to reactivate proteins at low protein concentrations (owing to their tendency to aggregate at high concentrations) was overcome by solubilization of inclusion bodies in alkaline solution and immobilization of proteins on a strong and resistant anion exchanger. This procedure has an inherent advantage of combining refolding and purification procedures in one step. The solubilization of the fusion protein in an alkaline reagent with the use of an anion exchanger resulted in considerable purification of the recombinant protein at a fairly high concentration. The protein was soluble under mild conditions and reacted with antibodies against the "native" papilloma virus.
Department of Biochemistry, Institute of Hematology and Blood Transfusion, Prague, Czech Republic.
J
J
Suttnar
J E
JE
Dyr
E
E
Hamsíková
J
J
Novák
V
V
Vonka
D000837:Anion Exchange Resins; D000906:Antibodies / Q000276:immunology / Q000302:isolation & purification; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000737:chemistry*; D006801:Humans; D015151:Immunoblotting; D002479:Inclusion Bodies / Q000737:chemistry*; D027383:Papillomaviridae / Q000235:genetics / Q000378:metabolism*; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification; D014760:Viral Fusion Proteins / Q000737:chemistry* / Q000276:immunology / Q000302:isolation & purification*; D014764:Viral Proteins / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification
D000837:Anion Exchange Resins; D000906:Antibodies; D011994:Recombinant Proteins; D014760:Viral Fusion Proteins; D014764:Viral Proteins