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Purification and Characterization of Paraoxonase 1 (PON1) from Swiss Black, Holstein, and Montofon Bovines

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Abstract

Paraoxonase 1 (PON1: EC 3.1.8.1) is a calcium-dependent enzyme associated with high-density lipoproteins (HDLs) and has a protective effect against oxidation of low-density lipoproteins (LDLs) in mammals. PON1 is the best-studied member of a family of enzymes called serum paraoxonases, or PONs, identified in mammals and other vertebrates as well as in invertebrates. PONs exhibit a range of important activities, including drug metabolism and detoxification of organophosphates such as nerve agents. This study reports, for the first time, purification and biochemical characterization of serum PON1 from different bovine breeds namely Swiss Black, Holstein, and Montofon. Bovine serum PON1s were purified using ammonium sulfate precipitation followed by Sepharose-4B-l-tyrosine-1-naphthylamine hydrophobic interaction chromatography. SDS–polyacrylamide gel electrophoresis of the purified enzymes indicates a single band with an apparent MW of 43 kDa. The purified enzymes had a specific activity of 10.78, 27.00, and 22.38 U/mg for Swiss Black, Holstein, and Montofon bovines, respectively. The overall purification rates of our method were 262.47-, 2,476.90-, and 538.06-fold for Swiss Black, Holstein, and Montofon bovines, respectively. Furthermore, using phenyl acetate as a substrate, we determined the K M and V max values of the purified enzymes, as 0.80 mM, 1428.5 U/ml for Swiss Black; 0.40 mM, 714.3 U/ml for Holstein; and 0.50 mM, 1,111.1 U/ml for Montofon bovine. The present study has revealed that there is no substantial difference in PON1 activities among the studied bovine breeds.

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Acknowledgments

This work was carried out in the Balikesir University Research Center of Applied Sciences (BURCAS).

Declaration of Interest

The authors declare that there are no conflicts of interest.

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Authors and Affiliations

  1. Faculty of Science and Letters, Department of Chemistry, Aksaray University, 68100, Aksaray, Turkey

    Mahmut Erzengin

  2. Faculty of Agriculture, Department of Agricultural Biotechnology, Süleyman Demirel University, 32260, Isparta, Turkey

    Dudu Demir

  3. Susurluk Technical Vocational School of Higher Education, Balikesir University, 10100, Balikesir, Turkey

    Mikail Arslan

  4. Faculty of Science and Letters, Department of Biology, Balikesir University, 10145, Balikesir, Turkey

    Selma Sinan

Authors
  1. Mahmut Erzengin
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  2. Dudu Demir
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Correspondence to Mahmut Erzengin.

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Erzengin, M., Demir, D., Arslan, M. et al. Purification and Characterization of Paraoxonase 1 (PON1) from Swiss Black, Holstein, and Montofon Bovines. Appl Biochem Biotechnol 173, 1597–1606 (2014). https://doi.org/10.1007/s12010-014-0931-1

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  • DOI: https://doi.org/10.1007/s12010-014-0931-1

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