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Immobilization of Ene Reductase in Polyvinyl Alcohol Hydrogel

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Abstract

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 °C, respectively. The entrapped ER showed 34.4-fold more thermal stability than that of the free ER at 30 °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.

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Data will be made available on request.

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Funding

This research was supported by the Scientific Research Unit of Cukurova University (Grant No. FBA-2017-8812).

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Authors and Affiliations

  1. Imamoglu Vocational School, Cukurova University, Adana, Turkey

    Dilek Alagöz

  2. Chemistry Department, Science and Letters Faculty, Cukurova University, Adana, Turkey

    Nazli Ece Varan & S. Seyhan Tükel

  3. Vocational School of Acigol, University of Nevsehir Haci Bektas Veli, Nevsehir, Turkey

    Ali Toprak

  4. Chemical Engineering Department, Ceyhan Engineering Faculty, Cukurova University, Adana, Turkey

    Deniz Yildirim

Authors
  1. Dilek Alagöz
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  2. Nazli Ece Varan
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  3. Ali Toprak
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  4. S. Seyhan Tükel
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  5. Deniz Yildirim
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Contributions

NEV and AT carried out the experiments. DY analyzed the data, planned the experiments. DA wrote the manuscript and SST supervised the experiments.

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Correspondence to Deniz Yildirim.

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Alagöz, D., Varan, N.E., Toprak, A. et al. Immobilization of Ene Reductase in Polyvinyl Alcohol Hydrogel. Protein J 41, 394–402 (2022). https://doi.org/10.1007/s10930-022-10059-4

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