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KMID : 0043320070300101302
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Archives of Pharmacal Research
2007 Volume.30 No. 10 p.1302 ~ p.1308
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Purification and Biochemical Properties of Phospholipase D (PLD57) from Streptomyces sp. CS-57
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Simkhada Jaya Ram
Cho Seung-Sik
Lee Hyo-Jung
Yoo Jin-Cheol
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Abstract
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Streptomyces sp. CS-57, which was isolated from Korean soil, was found to produce phospholipase D (PLD57) as an extracellular enzyme when cultured in medium containing 2% glucose, 1.5% yeast extract, 0.5% trypton, and 0.1% calcium carbonate at 28oC, and 160-rpm. PLD57 was purified using Sepharose CL-6B column chromatography, and DEAE-Sepharose CL-6B ion exchange column chromatography. The specific activity of the purified enzyme increased 6.7 fold with 3% recovery. The purified enzyme was then analyzed using 12% SDS-PAGE, which revealed that the molecular mass of the purified enzyme was 55 kDa. PLD57 showed both hydrolytic (H) and transphosphatidylation (T) activity, and the optimum temperatures of these activities were found to be 45oC and 35oC, respectively. Similarly, both of these activities were found to be optimal at a pH of 7.5. In addition, even after being heat treated at 45oC for up to 2 h, the enzyme activity remained at 100%, and the H-activity was found to be stable at a pH of 6 to 8. Further, enzyme activity occurred in the presence of EDTA, indicating that metal ions are not required for their activity, although some metal ions did marginally increase the activity. Enzyme activity also increased by 75% in the presence of Triton-X 100 at a concentration of 0.375 %; however, none of the other detergents evaluated in this study were found to enhance enzyme activity.
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KEYWORD
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Phospholipase D, Purification, Streptomyces, Hydrolytic activity, Transphosphatidylation activity
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