Immobilization of Ene Reductase in Polyvinyl Alcohol Hydrogel

Alagöz D., Varan N. E. , Toprak A., Tükel S. S. , Yıldırım D.

PROTEIN JOURNAL, 2022 (Peer-Reviewed Journal) identifier identifier identifier

  • Publication Type: Article / Article
  • Publication Date: 2022
  • Doi Number: 10.1007/s10930-022-10059-4
  • Journal Name: PROTEIN JOURNAL
  • Journal Indexes: Science Citation Index Expanded
  • Keywords: Ene-reductase, NADPH, Entrapment, Polyvinyl alcohol, 2-cyclohexen-1-one, ENZYME IMMOBILIZATION, ENOATE REDUCTASE, DESIGN, COIMMOBILIZATION, SUPPORTS, FORMATE, GEL


In this study, ene reductase (ER) was entrapped in polyvinyl alcohol hydrogel, adsorbed on montmorillonite and immobilized covalently on glutaraldehyde activated 3-aminopropyl-functionalized silica gel. Although protein recovery yields were at least 85% for adsorption and covalent immobilization, only the encapsulated ER showed activity. The activity of free and entrapped ER preparations was measured by following NADPH-dependent reduction of 2-cyclohexen-1-one. The both protein recovery and activity recovery yields were calculated as 100% when 1 mg protein was used for immobilization. The both free and entrapped ER preparations showed the same optimum pH and temperature as 7.0 and 30 degrees C, respectively. The entrapped ER showed 34.4-fold more thermal stability than that of the free ER at 30 degrees C. Michaelis-Menten constant and maximum velocity values were 0.25 mM and 1.2 U/mg protein, respectively for the free ER towards 2-cyclohexen-1-one. The corresponding values were 1.5 mM and 0.9 U/mg protein for the entrapped ER. The results of time-course reduction of 2-cyclohexen-1-one showed that the entrapped ER catalyzed the reaction as effectively as the free ER. The entrapped ER remained 85% of its initial activity after 10 reused cycles.