Abstract
Alkaloids, which are naturally occurring amines, are biosynthesized and accumulated in plant tissues such as roots, leaves, and stems. Many alkaloids have pharmacological effects on humans and animals. Cytochrome P450 (P450 or CYP) monooxygenases are known to play key roles in the biosynthesis of alkaloids in higher plants. A cDNA clone encoding a P450 protein consisting of 502 amino acids was isolated from Petunia hybrida. The deduced amino acid sequence of the cDNA clone showed a high level of similarity with the other P450 species in the CYP71D family; hence, this novel P450 was named CYP71D14. Among plant P450 species, CYP71D14 had 45.7% similarity in its amino acid sequence to CYP71D12, which is involved in the biosynthesis of the indole alkaloids vinblastine and vincristine. Expression of CYP71D14 mRNA in Petunia plants was examined by Northern blot analysis by using a full-length cDNA of CYP71D14 as a probe. CYP71D14 mRNA was expressed most abundantly in the roots.
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Abbreviations
- DEPC:
-
Diethyl pyrocarbonate
- DTT:
-
Dithiothreitol
- RT-PCR:
-
Reverse transcription polymerase chain reaction
- SDS:
-
Sodium dodecyl sulfate
- SRS-4:
-
Substrate recognition site 4
- SSC:
-
Saline sodium citrate
- SSPE:
-
Sodium chloride–sodium phosphate–EDTA
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Acknowledgments
We thank Dr. David R. Nelson (Committee for Standardized Cytochrome P450 Nomenclature) for his assistance in assigning the CYP71D14 sequence to the cytochrome P450 gene family. We acknowledge Dr. Hideo Ohkawa (Fukuyama University) and Dr. Kiyoharu Oono (Kobe University) for their comments on this study. This work was supported by Scientific Research Grant No. 11760066 to H. Imaishi from the Ministry of Education of Japan.
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Communicated by J. Sadowski.
The nucleotide sequence of CYP71D14 has been submitted to the DDBJ, EMBL, and GenBank nucleotide databases under the accession number AB028462.
An erratum to this article can be found at http://dx.doi.org/10.1007/s11738-008-0139-5
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Imaishi, H., Matushita, T. Cloning, expression in yeast, and functional characterization of CYP71D14, a root-specific cytochrome P450 from Petunia hybrida . Acta Physiol Plant 30, 333–338 (2008). https://doi.org/10.1007/s11738-007-0127-1
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DOI: https://doi.org/10.1007/s11738-007-0127-1