Epidithiol Formation by an Unprecedented Twin Carbon–Sulfur Lyase in the Gliotoxin Pathway

Scharf, Daniel H.; Chankhamjon, Pranatchareeya; Scherlach, Kirstin; Heinekamp, Thorsten; Roth, Martin; Brakhage, Axel A.; Hertweck, Christian

doi:10.1002/anie.201205041

Cited by 60 publications

(43 citation statements)

References 31 publications

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“…16–20 In addition a sixth gene encoding the O -methyltransferase GliM may catalyze the methylation of the free thiols. 18,36 Homologs of these six gliotoxin genes can be identified in the acetylaranotin ( 1 ) gene cluster in A. terreus (Table 1).…”

Section: Discussionmentioning

confidence: 99%

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Biosynthetic Pathway for the Epipolythiodioxopiperazine Acetylaranotin in Aspergillus terreus Revealed by Genome-Based Deletion Analysis

et al. 2013

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Epipolythiodioxopiperazines (ETPs) are a class of fungal secondary metabolites derived from diketopiperazines. Acetylaranotin belongs to one structural subgroup of ETPs characterized by the presence of a seven-membered 4,5-dihydrooxepine ring. Defining the genes involved in acetylaranotin biosynthesis should provide a means to increase production of these compounds and facilitate the engineering of second-generation molecules. The filamentous fungus Aspergillus terreus produces acetylaranotin and related natural products. Using targeted gene deletions, we have identified a cluster of nine genes including one nonribosomal peptide synthetase gene, ataP, which is required for acetylaranotin biosynthesis. Chemical analysis of the wild type and mutant strains enabled us to isolate seventeen natural products from the acetylaranotin biosynthesis pathway. Nine of the compounds identified in this study are previously not reported natural products. Our data allow us to propose a biosynthetic pathway for acetylaranotin and related natural products.

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Section: Discussionmentioning

confidence: 99%

“…In the next step, the putative dipeptidase GliJ removes the Glu residues and GliI acts as a C-S lyase to yield the epidithiol moiety, which is oxidized to the disulfide bridge by the sulfhydryl oxidase GliT. 16–20 …”

Section: Introductionmentioning

confidence: 99%

Biosynthetic Pathway for the Epipolythiodioxopiperazine Acetylaranotin in Aspergillus terreus Revealed by Genome-Based Deletion Analysis

et al. 2013

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“…The gliotoxin biosynthesis gene cluster has been identified in the human pathogen Aspergillus fumigatus , however, the exact sequence of steps in gliotoxin biosynthesis has remained unclear. 6 Various pathways for gliotoxin biosynthesis have been proposed by different groups using either knock-out approaches in A. fumigatus targeting gliP , 7 gliT , 8 gliZ , 9 gliG , 10, 11 gliH , 12 gliK 13 or gliI 14 genes and by biochemical approaches. However to date, the role of other genes in gliotoxin biosynthesis remains unclear.…”

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confidence: 99%

Reconstitution of the early steps of gliotoxin biosynthesis in Aspergillus nidulans reveals the role of the monooxygenase GliC

Chang

Chiang

Yeh

et al. 2013

Bioorganic & Medicinal Chemistry Letters

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The gliotoxin, a member of the epipolythiodioxopiperazine (ETP), has received considerable attention from the scientific community for its wide range of biological activity. Despite the identification of gliotoxin cluster, however, the sequence of steps in the gliotoxin biosynthesis has remained elusive. As an alternative to the gene knock out and biochemical approaches used so far, here we report using a heterologous expression approach to determine the sequence of the early steps of gliotoxin biosynthesis in A. nidulans. We identified the GliC, a monooxygenases that involved in the second step of gliotoxin biosynthesis pathway through the catalyzing the hydroxylation at the α-position of l-Phe.

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“…Although bisdethiobis(methylthio)gliotoxin (BmGT) (Fig. 1) has not been reported in most studies relating to gliT, gliA, gliK, or gliG (a glutathione S-transferase responsible for biosynthetic intermediate sulfurization via bis-glutathionylation) gene cluster deletion (4,5,7,13,15,18,19), this metabolite is readily detectable in culture supernatants of A. fumigatus (8)(9)(10). Moreover, endogenous dithiol gliotoxin [GT-(SH) 2 ] and exogenous gliotoxin can be converted to BmGT via a novel, S-adenosylmethionine (SAM)-dependent gliotoxin bis-thiomethyltransferase (GtmA) (10).…”

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Interplay between Gliotoxin Resistance, Secretion, and the Methyl/Methionine Cycle in Aspergillus fumigatus

et al. 2015

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Mechanistic studies on gliotoxin biosynthesis and self-protection in Aspergillus fumigatus, both of which require the gliotoxin oxidoreductase GliT, have revealed a rich landscape of highly novel biochemistries, yet key aspects of this complex molecular architecture remain obscure. Here we show that an A. fumigatus ⌬gliA strain is completely deficient in gliotoxin secretion but still retains the ability to efflux bisdethiobis(methylthio)gliotoxin (BmGT). This correlates with a significant increase in sensitivity to exogenous gliotoxin because gliotoxin trapped inside the cell leads to (i) activation of the gli cluster, as disabling gli cluster activation, via gliZ deletion, attenuates the sensitivity of an A. fumigatus ⌬gliT strain to gliotoxin, thus implicating cluster activation as a factor in gliotoxin sensitivity, and (ii) increased methylation activity due to excess substrate (dithiol gliotoxin) for the gliotoxin bis-thiomethyltransferase GtmA. Intracellular dithiol gliotoxin is oxidized by GliT and subsequently effluxed by GliA. In the absence of GliA, gliotoxin persists in the cell and is converted to BmGT, with levels significantly higher than those in the wild type. Similarly, in the ⌬gliT strain, gliotoxin oxidation is impeded, and methylation occurs unchecked, leading to significant S-adenosylmethionine (SAM) depletion and S-adenosylhomocysteine (SAH) overproduction. This in turn significantly contributes to the observed hypersensitivity of gliT-deficient A. fumigatus to gliotoxin. Our observations reveal a key role for GliT in preventing dysregulation of the methyl/methionine cycle to control intracellular SAM and SAH homeostasis during gliotoxin biosynthesis and exposure. Moreover, we reveal attenuated GliT abundance in the A. fumigatus ⌬gliK strain, but not the ⌬gliG strain, following exposure to gliotoxin, correlating with relative sensitivities. Overall, we illuminate new systems interactions that have evolved in gliotoxin-producing, compared to gliotoxin-naive, fungi to facilitate their cellular presence. Biosynthesis, self-protection mechanisms, and functionality of gliotoxin and related epidithiodiketopiperazine (ETP) molecular species, such as chaetocin and acetylaranotin, are attracting ever-increasing attention as a consequence of findings from highthroughput genome sequencing projects, application of gene deletion technologies, and mass spectrometric analytical methodologies (1-5). Indeed, existing paradigms of gliotoxin (Fig. 1) as a toxin and the perspective of the disulfide bridge-containing (oxidized) form as the final, or only, product are undergoing significant reconsideration (6-11).Self-protection against disulfide-containing metabolites appears to be essential in both fungi and bacteria. It has been demonstrated that the gliotoxin oxidoreductase GliT (12), encoded within the gli cluster, protects Aspergillus fumigatus against exogenous gliotoxin and is essential for gliotoxin biosynthesis (12, 13). A similar mechanism for self-protection against holomycin in Streptomyces clavulige...

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Epidithiol Formation by an Unprecedented Twin Carbon–Sulfur Lyase in the Gliotoxin Pathway

Cited by 60 publications

References 31 publications

Biosynthetic Pathway for the Epipolythiodioxopiperazine Acetylaranotin in Aspergillus terreus Revealed by Genome-Based Deletion Analysis

Biosynthetic Pathway for the Epipolythiodioxopiperazine Acetylaranotin in Aspergillus terreus Revealed by Genome-Based Deletion Analysis

Reconstitution of the early steps of gliotoxin biosynthesis in Aspergillus nidulans reveals the role of the monooxygenase GliC

Interplay between Gliotoxin Resistance, Secretion, and the Methyl/Methionine Cycle in Aspergillus fumigatus

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