Cytochrome P450 limonene hydroxylases of Mentha species.

The oxygenation pattern of the monoterpenoids of mint (Mentha) species is determined by regiospecific cytochrome P450-catalyzed hydroxylation of the common olefinic precursor (-)-limonene. In peppermint, C3-allylic hydroxylation leads to (-)-trans-isopiperitenol that ultimately is converted to (-)-menthol, whereas in spearmint, C6-allylic hydroxylation leads to (-)-trans-carveol that is oxidized to (-)-carvone. The limonene-6-hydroxylase and the cytochrome P450 reductase were purified from the oil glands of spearmint, and the system was reconstituted. Amino acid sequences from the purified hydroxylase were utilized to design primers with which a large, non-degenerate PCR product was prepared. This probe was employed to screen a spearmint oil gland cDNA library from which the corresponding full-length cDNA was isolated. This clone provides the tool for isolating the homologous cDNA species from peppermint and related Mentha species.