Publications

• Demonstration that menthofuran synthase of mint (Mentha) is a cytochrome P450 monooxygenase: cloning, functional expression, and characterization of the responsible gene.

  Bertea C.M., Schalk M., Karp F., Maffei M., Croteau R.

  (+)-Menthofuran is an undesirable monoterpenoid component of peppermint (Mentha x piperita) essential oil that is derived from the alpha,beta-unsaturated ketone (+)-pulegone. Microsomal preparations, from the oil gland secretory cells of a high (+)-menthofuran-producing chemotype of Mentha pulegiu ... >> More

  (+)-Menthofuran is an undesirable monoterpenoid component of peppermint (Mentha x piperita) essential oil that is derived from the alpha,beta-unsaturated ketone (+)-pulegone. Microsomal preparations, from the oil gland secretory cells of a high (+)-menthofuran-producing chemotype of Mentha pulegium, transform (+)-pulegone to (+)-menthofuran in the presence of NADPH and molecular oxygen, implying that menthofuran is synthesized by a mechanism analogous to that of mammalian liver cytochrome P450s involving the hydroxylation of the syn-methyl group of (+)-pulegone, spontaneous intramolecular cyclization to the hemiketal, and dehydration to the furan. An abundant cytochrome P450 clone from a peppermint oil gland cell cDNA library was functionally expressed in Saccharomyces cerevisiae and Escherichia coli and shown to encode the (+)-menthofuran synthase (i.e., (+)-pulegone-9-hydroxylase). The full-length cDNA contains 1479 nucleotides, and encodes a protein of 493 amino acid residues of molecular weight 55,360, which bears all of the anticipated primary structural elements of a cytochrome P450 and most closely resembles (35% identity) a cytochrome P450 monoterpene hydroxylase, (+)-limonene-3-hydroxylase, from the same source. The availability of this gene permits transgenic manipulation of peppermint to improve the quality of the derived essential oil. << Less

  Arch. Biochem. Biophys. 390:279-286(2001) [PubMed] [EuropePMC]

• Menthofuran regulates essential oil biosynthesis in peppermint by controlling a downstream monoterpene reductase.

  Mahmoud S.S., Croteau R.B.

  (+)-Pulegone is a central intermediate in the biosynthesis of (-)-menthol, the most significant component of peppermint essential oil. Depending on environmental conditions, this branch point metabolite may be reduced to (-)-menthone en route to menthol, by pulegone reductase (PR), or oxidized to ... >> More

  (+)-Pulegone is a central intermediate in the biosynthesis of (-)-menthol, the most significant component of peppermint essential oil. Depending on environmental conditions, this branch point metabolite may be reduced to (-)-menthone en route to menthol, by pulegone reductase (PR), or oxidized to (+)-menthofuran, by menthofuran synthase (MFS). To elucidate regulation of pulegone metabolism, we modified the expression of mfs under control of the CaMV 35S promoter in transformed peppermint plants. Overexpression and cosuppression of mfs resulted in the respective increase or decrease in the production of menthofuran, indicating that the control of MFS resides primarily at the level of transcription. Significantly, in both WT peppermint as well as in all transformed plants, the flux of (+)-pulegone through PR correlated negatively with the essential oil content of menthofuran, such that menthofuran, and pulegone increased, or decreased, in concert. These results suggested that menthofuran itself might influence the reduction of pulegone. Although (+)-menthofuran did not inhibit (+)-PR activity, stem feeding with menthofuran selectively decreased pr transcript levels in immature leaves, thereby accounting for decreased reductase activity and increased pulegone content. These data demonstrate that the metabolic fate of (+)-pulegone is controlled through transcriptional regulation of mfs and that menthofuran, either directly or indirectly, influences this process by down-regulating transcription from pr and/or decreasing pr message stability. The ability to reduce both menthofuran and pulegone levels is of commercial significance in improving essential oil quality; however, the physiological rationale for such complex regulation is presently unclear. << Less

  Proc. Natl. Acad. Sci. U.S.A. 100:14481-14486(2003) [PubMed] [EuropePMC]