Remarkable substrate-specificity of CYP6AB3 in Depressaria pastinacella, a highly specialized caterpillar

• Published in: Insect molecular biology Vol. 15; no. 2; pp. 169 - 179
• Main Authors: Mao, W, Rupasinghe, S, Zangerl, A.R, Schuler, M.A, Berenbaum, M.R
• Format: Journal Article
• Language: English
• Published: Oxford, UK Oxford, UK : Blackwell Publishing Ltd 01.04.2006; Blackwell Publishing Ltd
• Subjects: Animals; Apiaceae; binding properties; Binding Sites; catalytic site; Cell Line; complementary DNA; cytochrome P-450; Cytochrome P-450 Enzyme System - metabolism; cytochrome P-450 monooxygenase; cytochrome P450 monooxygenases; Depressaria pastinacella; enzyme activity; food plants; furanocoumarin metabolism; Furocoumarins - metabolism; gene expression; homology modelling; host specificity; Houseflies - enzymology; imperatorin; Larva - enzymology; larvae; metabolic detoxification; Models, Molecular; molecular models; Moths - enzymology; NADPH-Ferrihemoprotein Reductase - metabolism; plant-insect interactions; psoralens; Substrate Specificity
• ISSN: 0962-1075; 1365-2583
• DOI: 10.1111/j.1365-2583.2006.00623.x

The parsnip webworm, Depressaria pastinacella, a specialist on two genera in Apiaceae, feeds exclusively on the furanocoumarin-containing reproductive structures of its host plants. This caterpillar relies principally on cytochrome P450-mediated detoxification for coping with the high concentrations of furanocoumarins in its diet. A cDNA encoding the furanocoumarin-inducible P450 CYP6AB3 from this species was coexpressed with house-fly NADPH P450 reductase in baculovirus-infected Sf9 cells and tested for binding and metabolism of the six furanocoumarins typically encountered in host plant tissues. Only imperatorin and bergapten bind in close proximity to the catalytic haem and only imperatorin is metabolized (V[subscript max] and K[subscript m] of 2.412 pmol/min per pmol P450 and 94.28 [micro]m, respectively). Purification of the imperatorin metabolite by normal phase HPLC and characterization of its structure by MS-MS analysis indicate that CYP6AB3 initially epoxidizes the carbon-carbon pi-bond on the isoprenyl side chain on imperatorin. An improved molecular model for the CYP6AB3 protein based on this biochemical characterization and the recently defined mammalian CYP3A4 crystal structure provides insight into the remarkable substrate specificity of this protein.