N2 - Fungi are a rich source of bioactive secondary metabolites, and mushroom-forming fungi (Agaricomycetes) are especially known for the synthesis of numerous bioactive and often cytotoxic sesquiterpenoid secondary metabolites. Compared with the large number of sesquiterpene synthases identified in plants, less than a handful of unique sesquiterpene synthases have been described from fungi. Here we describe the functional characterization of six sesquiterpene synthases (Cop1 to Cop6) and two terpene-oxidizing cytochrome P450 monooxygenases (Cox1 and Cox2) from Coprinus cinereus. The genes were cloned and, except for cop5, functionally expressed in Escherichia coli and/or Saccharomyces cerevisiae. Cop1 and Cop2 each synthesize germacrene A as the major product. Cop3 was identified as an α-muurolene synthase, an enzyme that has not been described previously, while Cop4 synthesizes δ-cadinene as its major product. Cop6 was originally annotated as a trichodiene synthase homologue but instead was found to catalyse the highly specific synthesis of α-cuprenene. Coexpression of cop6 and the two monooxygenase genes next to it yields oxygenated α-cuprenene derivatives, including cuparophenol, suggesting that these genes encode the enzymes for the biosynthesis of antimicrobial quinone sesquiterpenoids (known as lagopodins) that were previously isolated from C. cinereus and other Coprinus species.
A new scheme for the biosynthesis of trichothecenes is proposed in which ITdiol and isotrichotriol are intermediates in the production of isotrichodermol from TDN.
Two novel compounds, 15-deacetyl-7,8-dihydroxycalonectrin (15-deacetylDHC) and 8a-hydroxyisotrichodiol were isolated from F.
Stereoselective synthesis of racemic trichodiene
We now report the results of incubation of trichodiene synthase with two analogues of the natural FPP substrate, 4-methylfarnesyl diphosphate (4M-FPP) and the less reactive substrate analogue 2-fluorofarnesyl diphosphate (2F-FPP) (). While cylization of FPP was found to give predominantly trichodiene, accompanied by trace amounts of several aberrant sesquiterpenes previously observed only with trichodiene synthase mutants, cyclization of both FPP analogues gave rise to complex mixtures of sesquiterpene analogues. When crystals of trichodiene synthase were soaked in a buffer solution containing 2FFPP, the resulting crystals were shown by X-ray crystallography to contain bound Mg2+ 3-PPi. We also report the crystal structure of the complex of trichodiene with Mg2+ 3-PPi and the benzyl triethylammonium cation (BTAC), the chloride salt of which is used as a phase transfer catalyst for synthetic organic chemistry; BTAC partially mimics the bisabolyl carbocation or other carbocation intermediates (). Taken together, these findings suggest that trichodiene synthase, and perhaps many terpene cyclases, may be capable of substantial versatility in binding and catalysis with unnatural substrates.
protein synthesis-inhibiting, trichothecene mycotoxins.
This review deals with the synthetic studies on trichothecane-type sesquiterpenes and verrucarins. Although more than forty compounds were isolated from natural sources, only a few examples of total synthesis of natural products in this class have been reported. They are trichodermin, trichodermol, 12, 13-epoxytrichothec-9-ene, and trichodiene. The total synthesis of an important sesquiterpene, verrucarol, having tow hydroxyl groups at C-4 and C-15 positions, has not yet succeeded. However, some synthetic studies towards the total synthesis of verrucarol are discussed. In the last part, partial syntheses of macrocyclic derivatives of verrucarol, tetrahydroverrucarin J and 2'-deoxy-3'-hydroxytetrahydroverrucarin A, are described.