Biosynthesis of a novel cyclic C35-terpene via the cyclisation of a Z-type C35-polyprenyl diphosphate obtained from a nonpathogenic Mycobacterium species

Abstract

Lipid components from 12 nonpathogenic Mycobacterium species were analysed. A novel cyclic C₃₅-terpene, named heptaprenylcycline 1, was obtained from 3 species, while octahydroheptaprenol 2, which has 3 Z-double bonds, was obtained from 6 species. The amounts of 1 and 2 in the cultured cells increased after the 4- to 6-d stationary phase. The yield of 1 was considerably greater at a higher temperature of 37 °C than at an optimal temperature of 28 °C, while that of 2 remained unchanged at all temperatures. A feeding experiment with D-[1-¹³C]glucose revealed that 1 was produced viaisopentenyl diphosphate, which is a metabolite of glycolysis and the methylerythritol phosphate pathway. The conversion of octahydroheptaprenyl diphosphate 2-PP to 1 was successful by using the cell-free extracts of M. chlorophenolicum, demonstrating that 2-PP is the biosynthetic intermediate of 1. This is the first example of the biosynthesis of a natural terpenevia the cyclisation of a linear C₃₅-isoprenoid. The substrate 2-PP for C₃₅-terpene cyclase has Z-type prenyl moieties; however, terpene cyclases usually employ E-type isoprenoids. The gene encoding the terpene cyclase that cyclises prenyl diphosphate containing Z-double bonds as the natural substrate has not yet been detected. Despite a careful search using the FASTA3 program, we could not detect any gene that is homologous to the known diphosphate-triggered type of mono-, sesqui- and diterpene cyclases in the genome of M. vanbaalenii, the DNA sequence of which has recently been elucidated. This suggests that a novel type of terpene cyclase might exist in the nonpathogenic Mycobacterium species.