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KMID : 0613820030130060879
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Journal of Life Science
2003 Volume.13 No. 6 p.879 ~ p.888
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Cloning of Cytochrome P450 Gene involved in the Pathway of Capsidiol Biosynthesis in Red Pepper Cells
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Kwon Soon-Tae
Kim Jae-Sung
Jung Do-Chul
Jeong Jeong-Hag
Hwang Jae-Moon
Oh Sei-Myoung
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Abstract
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In order to measure the enzyme activity of 5-epi-aristolochene hydroxylase, one of cytochrome P450 (P450) enzymes in eicitor-treated pepper cell, we used in vivo assay method and demonstrated a dramatic suppression of the activity by P450-inhibitors, ancymidol and ketocornazole. Using RT-PCR method with degenerate primer of the well conserved domains found within most P450-enzymes, and using cDNA library screening method, one distinct cDNA, being designated P450Hy01, was successfully isolated from elicitor-treated pepper cells. P450Hy01 mRNA was all induced in elicitor-treated cells whereas never induced in control cells. Moreover, levels of P450Hy01 expression were highly correlated with the levels of extracellular capsidiol production by different elicitors in cell cultures. P450Hy01 transcript was also induced by several other elicitors such as, cellulase, arachidonic acid, jasmonic acid, yeast extract as well as UV stress. P450Hy01 sequence contained high probability amino acid matches to known plant P450 genes and ORF with a conserved FxxGxRxCxG heme-binding domain. P450Hy01 cDNA showed 98% of homology in sequence of nucleotide as well as amino acid to 5-epi-aristolochene-1,3-hydroxylase(5EA1,3H) which has been isolated in tobacco cells, suggesting that P450Hy01 is prominent candidate gene for P450-enzyme encoding 5EA1,3H in pepper cell.
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KEYWORD
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red pepper, elicitor, heme-binding domain, cytochrome P450, 5-epi-aristolochene hydroxylase
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