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Biotechnology

Year: 2008 | Volume: 7 | Issue: 4 | Page No.: 751-756
Research Article

Purification and Characterization of Extra-Cellular Cholesterol Oxidase From Rhodococcus sp. PTCC 1633

M.T. Yazdi
Department of Pharmaceutical Biotechnology, Faculty of Pharmacy, Tehran University of Medical Sciences, P.O. Box 14155-6451, Tehran 14174, Iran
Z.T. Yazdi
Department of Pharmaceutical Biotechnology, Faculty of Pharmacy, Tehran University of Medical Sciences, P.O. Box 14155-6451, Tehran 14174, Iran
A. Ghasemian
Department of Pharmaceutical Biotechnology, Faculty of Pharmacy, Tehran University of Medical Sciences, P.O. Box 14155-6451, Tehran 14174, Iran
G. Zarrini
Department of Animal Biology, Faculty of Natural Sciences, University of Tabriz, P.O. Box 51666-16471, Tabriz 51664, Iran
N.H. Olyaee
Food and Drug Control Laboratories, P.O. Box 11136, Iran
Z. Sepehrizadeh
Department of Pharmaceutical Biotechnology, Faculty of Pharmacy, Tehran University of Medical Sciences, P.O. Box 14155-6451, Tehran 14174, Iran
In this study to isolate cholesterol oxidase (Cho) producing microorganism, a bacterium with high ability to produce Cho was found. It was identified with morphological, biochemical and molecular methods as a new species of Rhodococcus. Rhodococcus sp. PTCC 1633 has both extra-cellular and intracellular Cho. It was cultured in optimized condition and extra-cellular Cho concentrated by Three Phase Partitioning (TPP) with a recovery of 85%. The concentrated enzyme was purified by one step ion exchange chromatography. On SDS-PAGE the purified enzyme showed a molecular weight of about 55 kDa. The enzyme was active in a wide rang of pH and temperature with an optimum pH and temperature of 7.0-7.5 and 40°C, respectively, for activity. The Km value for this enzyme was 15 μM. A thermal stability experiment showed high stability at 40°C in 24 h.
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