A Role in 15-Deacetylcalonectrin Acetylation in the Non-Enzymatic Cyclization of an Earlier Bicyclic Intermediate in Trichothecene Biosynthesis.

The trichothecene biosynthesis in begins with the cyclization of farnesyl pyrophosphate to trichodiene, followed by subsequent oxygenation to isotrichotriol. This initial bicyclic intermediate is further cyclized to isotrichodermol (ITDmol), a tricyclic precursor with a toxic trichothecene skeleton. Although the first cyclization and subsequent oxygenation are catalyzed by enzymes encoded by and , the second cyclization occurs non-enzymatically. Following ITDmol formation, the enzymes encoded by , , , and catalyze 3--acetylation, 15-hydroxylation, 15--acetylation, and A-ring oxygenation, respectively. In this study, we extensively analyzed the metabolites of the corresponding pathway-blocked mutants of . The disruption of these genes, except , led to the accumulation of tricyclic trichothecenes as the main products: ITDmol due to disruption; a mixture of isotrichodermin (ITD), 7-hydroxyisotrichodermin (7-HIT), and 8-hydroxyisotrichodermin (8-HIT) due to disruption; and a mixture of calonectrin and 3-deacetylcalonectrin due to disruption. However, the Δ mutant accumulated substantial amounts of bicyclic metabolites, isotrichotriol and trichotriol, in addition to tricyclic 15-deacetylcalonectrin (15-deCAL). The ΔΔ double gene disruptant transformed ITD into 7-HIT, 8-HIT, and 15-deCAL. The deletion of and overexpression of and trichothecene regulatory genes did not result in the accumulation of 15-deCAL in the transgenic strain. Thus, the absence of Tri3p and/or the presence of a small amount of 15-deCAL adversely affected the non-enzymatic second cyclization and C-15 hydroxylation steps.