Affinity Thermoprecipitation of Yeast Alcohol Dehydrogenase through Metal Ion‐Promoted Binding with Eudragit‐Bound Cibacron Blue 3GA

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Abstract
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 °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.