Enzymatic synthesis of C40 carotenes by cell-free preparation from Halobacterium cutirubrum

Abstract

[14C]Mevalonate or [14C]isopentenyl pyrophosphate was found to be converted to trans-phytoene, trans-phytofluene, lycopene, and β-carotene by a cell-free 270 000 × g supernatant fraction prepared from Halobacterium cutirubrum cells that were broken by manual grinding with glass beads. Incubations were done under N2 in the dark at 37 °C in 4 M NaCl in presence of FAD, NADP, and MgCl2; ATP was also added when mevalonate was the substrate. This system was also capable of converting trans-[14C]phytoene to β-carotene via the intermediates trans-phytofluene, ζ-carotene, neurosporene, lycopene, and γ-carotene. Each of these labelled intermediates on incubation separately with the same enzyme system was shown to be converted to the intermediates farther down the pathway. The results of this study show that the biosynthetic pathway for the formation of C40 carotenes in H. cutirubrum proceeds as follows: isopentenyl pyrophosphate [Formula: see text] trans-phytoene → trans-phytofluene → ζ-carotene → neurosporene → lycopene → γ-carotene → β-carotene. This pathway differs from that in higher plants in that the cis isomers of phytoene and phytofluene are not on the main pathway of carotene biosynthesis, as they are in higher plants. Furthermore, trans-phytoene, which has not been reported to have any role in higher plants, appears to be the main intermediate in carotene biosynthesis in H. cutirubrum.