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Defense-Related Polyphenol Oxidase from Hevea brasiliensis Cell Suspension: Purification and Characterization

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Abstract

Polyphenol oxidase (PPO) was examined from the extract of leaf, seed, and cell suspension of Hevea brasiliensis, a rubber plant. The defense-related isozyme from Hevea cell suspension induced by culture filtrate of Phytophthora palmivora or by agitation stress was isolated through anion exchange and affinity chromatography, respectively. A 104-purification fold, migrated as a single band of 70 kDa on sodium dodecyl sulfate–polyacrylamide gel electrophoresis of PPO, was obtained after further purified by the preparative gel electrophoresis. Based on reaction with catechol and dopamine but not with p-cresol and guaiacol, it is a diphenol-type PPO. The values of V max /K m ratio indicated that catechol was the most specific substrate. The optimal activity of the purified PPO was observed at pH 6.0. The PPO activity was retained at pH 4.0–10.0 and temperature 10–60 °C. The inhibitors which completely inhibited the activity were ascorbic acid, dithiothreitol, and β-mercaptoethanol while sodium azide was a poor inhibitor. The PPO obtained from Hevea cell suspension possesses high specific activity and is stable at wide range of pH and temperature. It is therefore suitable for extreme condition uses and may lead to an alternative source of PPO in various industrial applications.

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Acknowledgments

This work was supported by the grants from Biochemistry Excellence Strengthening Program and by the Graduate Research Fund from Prince of Songkla University, Thailand to Ms. Nisaporn Muhamad. We thank Prof. Dr. Brian Hodgson and Mr. Michael Benjamin Lane for a generous revision of the manuscript and valuable comments.

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Authors and Affiliations

  1. Department of Biochemistry, Faculty of Science, Prince of Songkla University, Hat-Yai, Songkhla, 90112, Thailand

    Nisaporn Muhamad, Nion Chirapongsatonkul & Nunta Churngchow

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  1. Nisaporn Muhamad
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  2. Nion Chirapongsatonkul
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  3. Nunta Churngchow
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Correspondence to Nunta Churngchow.

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Muhamad, N., Chirapongsatonkul, N. & Churngchow, N. Defense-Related Polyphenol Oxidase from Hevea brasiliensis Cell Suspension: Purification and Characterization. Appl Biochem Biotechnol 167, 177–189 (2012). https://doi.org/10.1007/s12010-012-9690-z

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