One-Step Immobilization and Purification of His-Tagged Enzyme Using Poly(2-acetamidoacrylic acid) Hydrogel

다국어 초록 (Multilingual Abstract)

Ni2+-Complexed poly(2-acetamidoacrylic acid) (PAAA) hydrogel support was developed for the one-step immobilization and purification of recombinant histidine-tagged glutamyl aminopeptidase (His-tagged GAP). Ni2+-PAAA hydrogel was prepared from the polymerization of 2-acetamidoacrylic acid (AAA) and 2,2-[(1,4-dioxo-1,4-butanediyl) diamino] bis(2-propenoic acid) (DBDBPA) with potassium persulfate in dimethyl sulfoxide (DMSO),followed by Ni2+complexation. His-tagged GAP was immobilized directly from the cell lysate onto the Ni2+-PAAA hydrogel support and then purified. Catalytic activity of immobilized His-tagged GAP for the hydrolysis of alanylpara-nitroanilide revealed 90% conversion after 30 min of incubation, indicating sustained catalytic activity. The hydrogel-immobilized enzyme also exhibited enhanced thermal stability of sustained 70% activity after 1 h incubation at 60 °C, while the free enzyme activity was reduced to 50% at the same condition. After four cycles of hydrogel regeneration, the immobilized enzyme lost only 20% of its initial activity. Ni2+-PAAA hydrogel provided a new and convenient immobilization/purification system for His-tag enzymes through easy and simple procedures.

참고문헌 (Reference)

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