Cobalt-chelated magnetic particles for one-step purification and immobilization of His6-tagged Escherichia coli γ-glutamyltranspeptidase

Abstract

Cobalt-chelated magnetic (Fe3O4-AA-ANTA-Co2 +) particles were prepared and one-step purification and immobilization of His6-tagged Escherichia coli γ-glutamyltranspeptidase (His6-EcGGT) using these particles were evaluated. The optimal conditions for the adsorption of His6-EcGGT to Fe3O4-AA-ANTA-Co2 + particles were found to be 24.7 U g–1 adsorbent, pH 6.5, 300 mM NaCl and 30 min incubation at 4°C, while the elution solution was optimized to be 50 mM phosphate buffer (pH 6.5) containing 150 mM imidazole and 300 mM NaCl. The immobilized His6-EcGGT was recycled five times without significant loss of GGT activity. The average yield rate for the synthesis l-theanine from glutamine and ethylamine reached 56.7%. These results indicate that one-step affinity purification and immobilization of His6-EcGGT by Fe3O4-AA-ANTA-Co2 + particles might serve as an effective process for industrial application.