Fusarium Tri4 encodes a key multifunctional cytochrome P450 monooxygenase for four consecutive oxygenation steps in trichothecene biosynthesis

Tokai, Takeshi; Koshino, Hiroyuki; Takahashi-Ando, Naoko; Sato, Masayuki; Fujimura, Masatake; Kimura, Makoto

doi:10.1016/j.bbrc.2006.12.033

Cited by 121 publications

(100 citation statements)

References 22 publications

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“…Multifunctional cytochrome P450 enzymes that perform sequential oxidations have also been identified in other fungal secondary metabolism pathways including gibberellin biosynthesis in Fusarium fujikuroi (27,34) and trichothecene biosynthesis in Fusarium spp. (33). For the latter, an enzyme from Trichothecium roseum is able to perform only three of four oxidations performed by its Fusarium homolog, thus representing a loss or gain of function similar to what we are proposing for PaxQ/AtmQ.…”

Section: Discussionsupporting

confidence: 59%

Identification of Two Aflatrem Biosynthesis Gene Loci in Aspergillus flavus and Metabolic Engineering of Penicillium paxilli To Elucidate Their Function

Nicholson

Koulman

Monahan

et al. 2009

Appl Environ Microbiol

134

148

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Aflatrem is a potent tremorgenic toxin produced by the soil fungus Aspergillus flavus, and a member of a structurally diverse group of fungal secondary metabolites known as indole-diterpenes. Gene clusters for indole-diterpene biosynthesis have recently been described in several species of filamentous fungi. A search of Aspergillus complete genome sequence data identified putative aflatrem gene clusters in the genomes of A. flavus and Aspergillus oryzae. In both species the genes for aflatrem biosynthesis cluster at two discrete loci; the first, ATM1, is telomere proximal on chromosome 5 and contains a cluster of three genes, atmG, atmC, and atmM, and the second, ATM2, is telomere distal on chromosome 7 and contains five genes, atmD, atmQ, atmB, atmA, and atmP. Reverse transcriptase PCR in A. flavus demonstrated that aflatrem biosynthesis transcript levels increased with the onset of aflatrem production. Transfer of atmP and atmQ into Penicillium paxilli paxP and paxQ deletion mutants, known to accumulate paxilline intermediates paspaline and 13-desoxypaxilline, respectively, showed that AtmP is a functional homolog of PaxP and that AtmQ utilizes 13-desoxypaxilline as a substrate to synthesize aflatrem pathway-specific intermediates, paspalicine and paspalinine. We propose a scheme for aflatrem biosynthesis in A. flavus based on these reconstitution experiments in P. paxilli and identification of putative intermediates in wild-type cultures of A. flavus.

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Section: Discussionsupporting

confidence: 59%

Identification of Two Aflatrem Biosynthesis Gene Loci in Aspergillus flavus and Metabolic Engineering of Penicillium paxilli To Elucidate Their Function

Nicholson

Koulman

Monahan

et al. 2009

Appl Environ Microbiol

134

148

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“…This result has important implications for metabolic engineering of crop plants for enhanced disease resistance and also for the generation of antimicrobial triterpenes for other applications. Multifunctional P450 enzymes from other P450 families that catalyze both hydroxylation and epoxidation reactions have been described recently from bacteria (47)(48)(49)(50)(51) and fungi (52,53). Examples of both possible reaction orders (hydroxylation followed by epoxidation and epoxidation followed by hydroxylation) have been reported (Fig.…”

Section: Resultsmentioning

confidence: 99%

Biochemical analysis of a multifunctional cytochrome P450 (CYP51) enzyme required for synthesis of antimicrobial triterpenes in plants

Geisler

Hughes

Sainsbury

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

151

134

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Members of the cytochromes P450 superfamily (P450s) catalyze a huge variety of oxidation reactions in microbes and higher organisms. Most P450 families are highly divergent, but in contrast the cytochrome P450 14α-sterol demethylase (CYP51) family is one of the most ancient and conserved, catalyzing sterol 14α-demethylase reactions required for essential sterol synthesis across the fungal, animal, and plant kingdoms. Oats (Avena spp.) produce antimicrobial compounds, avenacins, that provide protection against disease. Avenacins are synthesized from the simple triterpene, β-amyrin. Previously we identified a gene encoding a member of the CYP51 family of cytochromes P450, AsCyp51H10 (also known as Saponin-deficient 2, Sad2), that is required for avenacin synthesis in a forward screen for avenacin-deficient oat mutants. sad2 mutants accumulate β-amyrin, suggesting that they are blocked early in the pathway. Here, using a transient plant expression system, we show that AsCYP51H10 is a multifunctional P450 capable of modifying both the C and D rings of the pentacyclic triterpene scaffold to give 12,13β-epoxy-3β,16β-dihydroxy-oleanane (12,13β-epoxy-16β-hydroxy-β-amyrin). Molecular modeling and docking experiments indicate that C16 hydroxylation is likely to precede C12,13 epoxidation. Our computational modeling, in combination with analysis of a suite of sad2 mutants, provides insights into the unusual catalytic behavior of AsCYP51H10 and its active site mutants. Fungal bioassays show that the C12,13 epoxy group is an important determinant of antifungal activity. Accordingly, the oat AsCYP51H10 enzyme has been recruited from primary metabolism and has acquired a different function compared to other characterized members of the plant CYP51 family-as a multifunctional stereo-and regio-specific hydroxylase in plant specialized metabolism.

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“…This series of possible biosynthetic steps indicates the role of Fsr3 as a multifunctional monooxygenase. So far, multiple oxidization reactions by a single enzyme have been described only for cytochrome P450 monooxygenases, including three of the four P450 monooxygenases in the gibberellin biosynthesis of F. fujikuroi (62,76,77) and Tri4 in the trichothecene biosynthesis of F. graminearum (73). In A. fumigatus, chain shortening of YWA1 was shown to be catalyzed by Ayg1, resulting in the formation of 1,3,6,8-tetrahydroxynaphthalene (T4HN) (27).…”

Section: Discussionmentioning

confidence: 99%

Biosynthesis of Fusarubins Accounts for Pigmentation of Fusarium fujikuroi Perithecia

Studt

Wiemann

Kleigrewe

et al. 2012

Appl Environ Microbiol

175

189

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ABSTRACTFusarium fujikuroiproduces a variety of secondary metabolites, of which polyketides form the most diverse group. Among these are the highly pigmented naphthoquinones, which have been shown to possess different functional properties for the fungus. A group of naphthoquinones, polyketides related to fusarubin, were identified inFusariumspp. more than 60 years ago, but neither the genes responsible for their formation nor their biological function has been discovered to date. In addition, although it is known that the sexual fruiting bodies in which the progeny of the fungus develops are darkly colored by a polyketide synthase (PKS)-derived pigment, the structure of this pigment has never been elucidated. Here we present data that link the fusarubin-type polyketides to a defined gene cluster, which we designatefsr, and demonstrate that the fusarubins are the pigments responsible for the coloration of the perithecia. We studied their regulation and the function of the single genes within the cluster by a combination of gene replacements and overexpression of the PKS-encoding gene, and we present a model for the biosynthetic pathway of the fusarubins based on these data.

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Fusarium Tri4 encodes a key multifunctional cytochrome P450 monooxygenase for four consecutive oxygenation steps in trichothecene biosynthesis

Cited by 121 publications

References 22 publications

Identification of Two Aflatrem Biosynthesis Gene Loci in Aspergillus flavus and Metabolic Engineering of Penicillium paxilli To Elucidate Their Function

Identification of Two Aflatrem Biosynthesis Gene Loci in Aspergillus flavus and Metabolic Engineering of Penicillium paxilli To Elucidate Their Function

Biochemical analysis of a multifunctional cytochrome P450 (CYP51) enzyme required for synthesis of antimicrobial triterpenes in plants

Biosynthesis of Fusarubins Accounts for Pigmentation of Fusarium fujikuroi Perithecia

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