Polyketide folding in higher plants: biosynthesis of the phenylanthraquinone knipholone.

The biosynthesis of knipholone, as an axially chiral phenylanthraquinone, in higher plants was examined by feeding experiments with [13C2]-labeled precursors. [13C2]-Acetate and advanced synthetic intermediates were fed to sterile cultures of Kniphofia pumila (Asphodelaceae), with subsequent NMR analysis on the isolated natural product involving 2D INADEQUATE and SELINQUATE experiments. Due to its uneven number of carbon atoms, and because of its uncertain decarboxylation site, the "northern" part of the molecule (i.e., the chrysophanol portion) might originate from four different cyclization modes. According to the labeling pattern of the product isolated after incorporation, this anthraquinone part of knipholone is formed by the so-called F folding mode (originally established for fungi). The acetophenone part of the molecule, which does not undergo a decarboxylation reaction, originates from four acetate units. The surprising lack of randomization of the intact [13C2] units in this "southern" part reveals the absence of a free symmetric intermediate as initially anticipated. This is in agreement with the intact incorporation of the "authentic" southern molecular portion, 4,6-dihydroxy-2-methoxyacetophenone, while the corresponding symmetrical candidate trihydroxyacetophenone was clearly not incorporated, showing that the O-methylation of the freshly cyclized tetraketide is the step that prevents symmetrization of the acetophenone.