Metabolic engineering of monoterpene biosynthesis: two-step production of (+)-trans-isopiperitenol by tobacco.
Monoterpenoid biosynthesis in tobacco was modified by introducing two subsequent enzymatic activities targeted to different cell compartments. A limonene-3-hydroxylase (lim3h) cDNA was isolated from Mentha spicata L. 'Crispa'. This cDNA was used to re-transform a transgenic Nicotiana tabacum'Petit Havana' SR1 (tobacco) line expressing three Citrus limon L. Burm. f. (lemon) monoterpene synthases producing (+)-limonene, gamma-terpinene and (-)-beta-pinene as their main products. The targeting sequences of these synthases indicate that they are probably localized in the plastids, whereas the sequence information of the P450 hydroxylase indicates targeting to the endoplasmatic reticulum. Despite the different location of the enzymes, the introduced P450 hydroxylase proved to be functional in the transgenic plants as it hydroxylated (+)-limonene, resulting in the emission of (+)-trans-isopiperitenol. Some further modifications of the (+)-trans-isopiperitenol were also detected, resulting in the additional emission of 1,3,8-p-menthatriene, 1,5,8-p-menthatriene, p-cymene and isopiperitenone.
Lücker
Joost
J
Business Unit Bioscience, Plant Research International, PO Box 16, 6700 AA Wageningen, The Netherlands.
Schwab
Wilfried
W
Franssen
Maurice C R
MC
Van Der Plas
Linus H W
LH
Bouwmeester
Harro J
HJ
Verhoeven
Harrie A
HA
D000595:Amino Acid Sequence; D002957:Citrus / Q000201:enzymology / Q000235:genetics; D003577:Cytochrome P-450 Enzyme System / Q000138:chemical synthesis / Q000235:genetics / Q000378:metabolism; D035264:Flowers / Q000201:enzymology; D008401:Gas Chromatography-Mass Spectrometry; D020868:Gene Silencing; D005822:Genetic Vectors; D031337:Mentha spicata / Q000235:genetics / Q000378:metabolism; D006899:Mixed Function Oxygenases / Q000138:chemical synthesis / Q000235:genetics; D008956:Models, Chemical; D008969:Molecular Sequence Data; D039821:Monoterpenes / Q000737:chemistry / Q000378:metabolism*; D030821:Plants, Genetically Modified; D013729:Terpenes / Q000737:chemistry / Q000378:metabolism*; D014026:Nicotiana / Q000235:genetics* / Q000378:metabolism; D014170:Transformation, Genetic; D014835:Volatilization
D039821:Monoterpenes; D013729:Terpenes; C044945:isopiperitenol; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; C120915:limonene-3-hydroxylase