Development of an expanded bed technique for an affinity purification of G6PDH from unclarified yeast cell homogenates

摘要: The use of an expanded bed STREAMLINE Red H-7B for the purification intracellular glycolytic enzyme glucose 6-phosphate dehydrogenase (G6PDH) directly from untreated preparations disrupted yeast cells has been investigated. Small-scale experiments, carried out in packed beds, have shown that optimal pH adsorption is 6.0 and enabled optimization elution conditions using a series eluents. dynamic capacity adsorbent G6PDH was determined small to be 28 units/mL. These results were used develop preparative scale separation 50 column. purified unclarified homogenate 99% yield with average factor eluted fraction 103. Cleaning-in-place (CIP) procedures 0.5 M NaOH 4M urea 60% (v/v) ethanol demonstrated can regenerated no loss or alteration expansion characteristics after many cycles operation.