A Multifunctional Monooxygenase XanO4 Catalyzes Xanthone Formation in Xantholipin Biosynthesis via a Cryptic Demethoxylation

Abstract

Xantholipin and several related polycyclic xanthone antibiotics feature a unique xanthone ring nucleus within a highly oxygenated, angular, fused hexacyclic system. In this study, we demonstrated that a flavin-dependent monooxygenase (FMO) XanO4 catalyzes the oxidative transformation of an anthraquinone to a xanthone system during the biosynthesis of xantholipin. In vitro isotopic labeling experiments showed that the reaction involves sequential insertion of two oxygen atoms, accompanied by an unexpected cryptic demethoxylation reaction. Moreover, characterizations of homologous FMOs of XanO4 suggested the generality of the XanO4-like-mediated reaction for the assembly of a xanthone ring in the biosynthesis of polycyclic xanthone antibiotics. These findings not only expand the repertoire of FMO activities but also reveal a novel mechanism for xanthone ring formation.

Sweating the assets of flavin cofactors: new insight of chemical versatility from knowledge of structure and mechanism

“…These include simple oxidations of thiols by oxidases to form thiocarbonyls [35], thereby extending the chemistry of oxidases beyond oxidation of C-O, C-C and C-N bonds. Furthermore, complex cascade chemistry has been postulated to occur in a flavin dependent decarboxylase-dehydrogenase-monooxygenase during assembly of the warhead a,b-epoxyketone proteasome inhibitors [36] and for the multifunctional monooxygenase XanO4 that catalyzes sequential insertion of two oxygen atoms and a cryptic demethoxylation in the biosynthesis of xantholipids [37]. These intriguing reactions extend further the already large family of biocatalysts represented by the flavin-dependent monooxygenases [22].…”

Biosynthesis of the pyrrolidine protein synthesis inhibitor anisomycin involves novel gene ensemble and cryptic biosynthetic steps

“…Several cryptic reactions in other natural product biosynthetic pathways have been reported recently. Cryptic methyl esterification (22), chlorination (23), acylation (24), and demethoxylation (25,26) have been shown to be indispensable in the biosynthesis of streptonigrin, coronatine, saframycin, and xantholipin, respectively. However, to the best of our knowledge, glucose, which is the main carbon source, is used to mediate the biosynthesis of natural products, has not been previously reported in microbial natural product pathways.…”

New insights into bacterial type II polyketide biosynthesis

“…The further isotope labeling experiments and high-resolution tandem mass spectrometry of proteinase K-digested EncM provide direct evidence for Fl N5[O] species, which may be derived from the direct reaction of O 2 with the flavin: hydrogen transfer from Fl red to O 2 , resulting in the production of flavin SQ and protonated superoxide, followed by the formation of a flavin-N5-peroxide via radical coupling of the formed protonated superoxide and anionic flavin semiquinone, and the subsequent water elimination affords the Fl N5[O] cofactor ( Figure 3A ) 12 . Recently, another flavin-dependent monooxygenase (MO), XanO4, was demonstrated to catalyze epoxidation and, through Baeyer-Villiger (BV) dual reaction, transform an anthraquinone into a xanthone skeleton during the biosynthesis of xantholipin 13 . Significantly, homologous comparisons indicated that the XanO4-mediated reaction was supposed to be the general model of constructing a xanthone ring in polycyclic xanthone antibiotic biosynthesis ( Figure 3B ).…”