Affinity thermoprecipitation of yeast alcohol dehydrogenase through metal ion-promoted binding with Eudragit-bound Cibacron blue 3GA.
Metal ion-promoted binding of Saccharomyces cerevisiae alcohol dehydrogenase to Cibacron Blue 3GA was used for its isolation by affinity precipitation with Eudragit-bound dye. The yeast cells were broken and the cell debris was separated by flocculation with poly(ethylene imine). The supernatant containing the enzyme activity was mixed with Eudragit--Cibacron Blue in the presence of zinc ions. The precipitation of the affinity complex was induced by the addition of 50 mM CaCl2 and a subsequent increase in temperature to 40 degrees C. The enzyme was desorbed by treating the precipitate with iminodiacetic acid solution. The procedure resulted in about 66% recovery of enzyme activity with more than 20-fold purification. Recycling of Eudragit--dye for enzyme purification was also shown to be possible.
Guoqiang
D
D
Department of Biotechnology, Lund University, Sweden.
Benhura
M A
MA
Kaul
R
R
Mattiasson
B
B
D000180:Acrylic Resins / Q000378:metabolism*; D000426:Alcohol Dehydrogenase / Q000302:isolation & purification* / Q000378:metabolism; D011232:Chemical Precipitation; D004396:Coloring Agents / Q000378:metabolism*; D008670:Metals; D011109:Polymethacrylic Acids; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D013057:Spectrum Analysis; D013379:Substrate Specificity; D014227:Triazines / Q000378:metabolism*; D015032:Zinc
D000180:Acrylic Resins; D004396:Coloring Agents; D008670:Metals; D011109:Polymethacrylic Acids; D014227:Triazines; C038300:methylmethacrylate-methacrylic acid copolymer; C012842:Cibacron Blue F 3GA; D000426:Alcohol Dehydrogenase; D015032:Zinc