Separation of enzymes from Candida boidinii crude extract by continuous flow zone electrophoresis.
Separation of the enzymes formate dehydrogenase, formaldehyde dehydrogenase and methanol oxidase from Candida boidinii crude extract has been explored using continuous flow zone electrophoresis in the VaP-22 and the scaled-up VaP-220 electrophoresis apparatus. Yields up to 95% and purification factors between 3 and 7 were obtained, together with separation of cell debris from the enzymes. Multiple injections of sample were used to obtain a protein throughput of 46.2 mg/h in the VaP-22. A tenfold higher throughput was achieved using the VaP-220. Correlation of the electrophoretic mobility in continuous flow zone electrophoresis with the elution behavior in ion-exchange chromatography confirmed the primary role of net surface charge in the separation of biological molecules. Proteins and enzymes with differences greater than 0.05 M elution molarities in ion-exchange chromatography can be separated. This corresponds to a preparative scale (mg/h or g/h) separation of proteins and enzymes whose difference in apparent electrophoretic mobility is greater than 0.70 x 10(-5) cm2/(V.s).
Nath
S
S
GBF-Gesellschaft für Biotechnologische, Forschung mbH, Braunschweig, FRG.
Schütte
H
H
Weber
G
G
Hustedt
H
H
Deckwer
W D
WD
D000429:Alcohol Oxidoreductases / Q000302:isolation & purification; D000445:Aldehyde Oxidoreductases / Q000302:isolation & purification; D002175:Candida / Q000201:enzymology*; D002852:Chromatography, Ion Exchange; D004586:Electrophoresis; D005560:Formate Dehydrogenases / Q000302:isolation & purification; D010088:Oxidoreductases / Q000302:isolation & purification*
D010088:Oxidoreductases; D000429:Alcohol Oxidoreductases; C020145:formaldehyde dehydrogenase (glutathione); C029293:methanol oxidase; D005560:Formate Dehydrogenases; D000445:Aldehyde Oxidoreductases