Evaluation of immobilized metal affinity chromatography for purification of penicillin acylase.
The aim of this work was to test immobilized metal affinity chromatography (IMAC) for the purification of penicillin acylase. After evaluation of different metals, Cu2+ was selected. Different samples were tested: pure penicillin acylase, industrial clarified feedstock and crude extract. After comparing two eluents, NH4Cl and imidazole, it appeared that although both gave good results for recovery and activity, NH4Cl was a more selective eluent with a higher fold purification than imidazole (4.64 versus 2.04). Moreover, we shown that a multistep gradient of NH4Cl, greatly increased the degree of purification (12.36) compared with the one-step process as control (4.64). In addition, good recovery was obtained (97-100%).
Fitton
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Ecole Supérieure de Technologie des Biomolécules de Bordeaux (ESTBB), UMR 5544, Université Victor Segalen Bordeaux 2, France.
Santarelli
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D000821:Animal Feed / Q000032:analysis; D002413:Cations, Divalent; D002846:Chromatography, Affinity / Q000379:methods*; D003300:Copper; D004926:Escherichia coli; D008670:Metals; D009532:Nickel; D010399:Penicillin Amidase / Q000302:isolation & purification*; D011485:Protein Binding; D015032:Zinc
D002413:Cations, Divalent; D008670:Metals; D003300:Copper; D009532:Nickel; D010399:Penicillin Amidase; D015032:Zinc