Efficient two-step chromatographic purification of penicillin acylase fromclarified Escherichia coli ultrasonic homogenate

A two-step chromatographic purification procedure from clarified Escherichi a coli ultrasonic homogenate was evaluated. The capture step included immob ilized metal affinity chromatography with Cu2+ as metal ion. Two elution me thods were performed: 1 M NH4Cl and 0.01 M imidazole. Respectively, we obta ined a different purification fold (16.5 to 3.15) and a similar result for the recovery of activity (90-99%). The best elution method was chosen for t he procedure. The second step, hydrophobic interaction chromatography, gave a 3.8-fold purification with 77.7% of activity. The total procedure gave a 66-fold purification in relation to the initial crude extract with 70% for the recovery of activity and was performed without any conditioning step a nd at the same pH value. (C) 2001 Elsevier Science B.V. All rights reserved .