Abstract:Gliotoxin (GT) is a mycotoxin produced by some species of ascomycete fungi
including the opportunistic human pathogen Aspergillus
fumigatus. In order to produce GT the host organism needs to have
evolved a self-protection mechanism. GT contains a redox-cycling disulfide
bridge that is important in mediating toxicity. Recently is has been
demonstrated that A. fumigatus possesses a novel
thiomethyltransferase protein called GtmA that has the ability to thiomethylate
GT in vivo, which aids the organism in regulat… Show more
“…Figure 6.( a ) Aspergillus fumigatus Δ gliT responds to exogenous gliotoxin by effecting excessive bis -thiomethylation on this metabolite, resulting in cellular SAM depletion. ( b ) The highly sensitive A. fumigatus Δg liT ::Δ gtmA cannot remove intracellular accumulated dithiol gliotoxin by S -methylation [6,35] or oxidation. This results in cell death as a result of the combined effect of gliotoxin toxicity and gli -cluster over-activation owing to the sustained presence of dithiol gliotoxin.…”
Section: Discussionmentioning
confidence: 99%
“…( b ) The highly sensitive A. fumigatus Δg liT ::Δ gtmA cannot remove intracellular accumulated dithiol gliotoxin by S -methylation [6,35] or oxidation. This results in cell death as a result of the combined effect of gliotoxin toxicity and gli -cluster over-activation owing to the sustained presence of dithiol gliotoxin.…”
Gliotoxin is an epipolythiodioxopiperazine (ETP) class toxin, contains a disulfide bridge that mediates its toxic effects via redox cycling and is produced by the opportunistic fungal pathogen Aspergillus fumigatus. Self-resistance against gliotoxin is effected by the gliotoxin oxidase GliT, and attenuation of gliotoxin biosynthesis is catalysed by gliotoxin S-methyltransferase GtmA. Here we describe the X-ray crystal structures of GtmA-apo (1.66 Å), GtmA complexed to S-adenosylhomocysteine (1.33 Å) and GtmA complexed to S-adenosylmethionine (2.28 Å), providing mechanistic insights into this important biotransformation. We further reveal that simultaneous elimination of the ability of A. fumigatus to dissipate highly reactive dithiol gliotoxin, via deletion of GliT and GtmA, results in the most significant hypersensitivity to exogenous gliotoxin observed to date. Indeed, quantitative proteomic analysis of ΔgliT::ΔgtmA reveals an uncontrolled over-activation of the gli-cluster upon gliotoxin exposure. The data presented herein reveal, for the first time, the extreme risk associated with intracellular dithiol gliotoxin biosynthesis—in the absence of an efficient dismutation capacity. Significantly, a previously concealed protective role for GtmA and functionality of ETP bis-thiomethylation as an ancestral protection strategy against dithiol compounds is now evident.
“…Figure 6.( a ) Aspergillus fumigatus Δ gliT responds to exogenous gliotoxin by effecting excessive bis -thiomethylation on this metabolite, resulting in cellular SAM depletion. ( b ) The highly sensitive A. fumigatus Δg liT ::Δ gtmA cannot remove intracellular accumulated dithiol gliotoxin by S -methylation [6,35] or oxidation. This results in cell death as a result of the combined effect of gliotoxin toxicity and gli -cluster over-activation owing to the sustained presence of dithiol gliotoxin.…”
Section: Discussionmentioning
confidence: 99%
“…( b ) The highly sensitive A. fumigatus Δg liT ::Δ gtmA cannot remove intracellular accumulated dithiol gliotoxin by S -methylation [6,35] or oxidation. This results in cell death as a result of the combined effect of gliotoxin toxicity and gli -cluster over-activation owing to the sustained presence of dithiol gliotoxin.…”
Gliotoxin is an epipolythiodioxopiperazine (ETP) class toxin, contains a disulfide bridge that mediates its toxic effects via redox cycling and is produced by the opportunistic fungal pathogen Aspergillus fumigatus. Self-resistance against gliotoxin is effected by the gliotoxin oxidase GliT, and attenuation of gliotoxin biosynthesis is catalysed by gliotoxin S-methyltransferase GtmA. Here we describe the X-ray crystal structures of GtmA-apo (1.66 Å), GtmA complexed to S-adenosylhomocysteine (1.33 Å) and GtmA complexed to S-adenosylmethionine (2.28 Å), providing mechanistic insights into this important biotransformation. We further reveal that simultaneous elimination of the ability of A. fumigatus to dissipate highly reactive dithiol gliotoxin, via deletion of GliT and GtmA, results in the most significant hypersensitivity to exogenous gliotoxin observed to date. Indeed, quantitative proteomic analysis of ΔgliT::ΔgtmA reveals an uncontrolled over-activation of the gli-cluster upon gliotoxin exposure. The data presented herein reveal, for the first time, the extreme risk associated with intracellular dithiol gliotoxin biosynthesis—in the absence of an efficient dismutation capacity. Significantly, a previously concealed protective role for GtmA and functionality of ETP bis-thiomethylation as an ancestral protection strategy against dithiol compounds is now evident.
“…31 The organic extracts were subsequently dried down and resuspended in methanol and analysed for gliotoxin content using RP-HPLC with UV detection (Shimadzu), using polar C18 RP-HPLC column (Phenomenex polar C18 Luna Omega column (150 mm  4.6 mm, 5 mm)) at a flow rate of 1 ml min…”
Section: Extraction and Measurement Of Extracellular And Intracellulamentioning
Significance to metallomicsDithiol gliotoxin is a near-terminal biosynthetic intermediate from the gliotoxin biosynthetic pathway in the human pathogen Aspergillus fumigatus. Chemically reduced gliotoxin, dithiol gliotoxin (DTG), is revealed as a biological zinc chelator, and conversely, zinc can relieve the hitherto cryptic fungal autotoxicity of DTG. There is a systems-wide impact of zinc chelation by DTG on the fungal proteome, and we suggest it is DTG, as opposed to gliotoxin, which chelates zinc from metalloproteins. Since gliotoxin can be sequestered by both fungi and bacteria, our findings infer a new avenue to interfere with, and exploit, cellular zinc homeostasis in microorganisms.
IntroductionGliotoxin and holomycin are microbial natural products, which are produced by fungal and bacterial spp., respectively (Fig. 1). [1][2][3] Both are low molecular mass metabolites, and each contains a disulphide bridge formed by the action of oxidoreductases, namely GliT and HlmI, on the respective dithiol precursor (Fig. 1). [4][5][6] Disulphide bridge formation is essential for microbial self-protection against these reactive dithiol intermediates, and is a pre-requisite for the Major Facilitator Superfamily transporter GliA-mediated secretion of gliotoxin by Aspergillus fumigatus.
5-8Both metabolites are also present as bis-thiomethylated forms, and gliotoxin bis-thiomethyltransferase GtmA converts dithiol gliotoxin (DTG) to bis-dethiobis(methylthio)gliotoxin (BmGT) in A. fumigatus, whereas the origin of the cognate activity against dithiol holomycin in Streptomyces clavuligeris is unknown (Fig. 1).
9,10Bernardo et al. have shown that upon uptake by eukaryotic cells,
“…Gliotoxin (GT), an alkaloid with a molecular mass of 326 Da, is the most important and well-known epipolythiodioxypipeazine (ETP)-type mycotoxin with biological active internal disulfide bridge (Smith et al, 2016). Since it was discovered by the 1930s, GT has been isolated from various fungal species, including Trichoderma, Aspergillus fumigatus, Eurotium chevalieri, Neosartorya pseudofischeri, some Penicillium spp., and Acremonium spp.…”
Gliotoxin (GT) is a fungal secondary metabolite that has attracted great interest due to its high biological activity since it was discovered by the 1930s. It exhibits a unique structure that contains a N-C = O group as the characteristics of the classical PSII inhibitor. However, GT's phytotoxicity, herbicidal activity and primary action targets in plants remain hidden. Here, it is found that GT can cause brown or white leaf spot of various monocotyledonous and dicotyledonous plants, being regarded as a potential herbicidal agent. The multiple sites of GT action are located in two photosystems. GT decreases the rate of oxygen evolution of PSII with an I 50 value of 60 µM. Chlorophyll fluorescence data from Chlamydomonas reinhardtii cells and spinach thylakoids implicate that GT affects both PSII electron transport at the acceptor side and the reduction rate of PSI end electron acceptors' pool. The major direct action target of GT is the plastoquinone Q B-site of the D1 protein in PSII, where GT inserts in the Q B binding niche by replacing native plastoquinone (PQ) and then interrupts electron flow beyond plastoquinone Q A. This leads to severe inactivation of PSII RCs and a significant decrease of PSII overall photosynthetic activity. Based on the simulated modeling of GT docking to the D1 protein of spinach, it is proposed that GT binds to the-Q B-site through two hydrogen bonds between GT and D1-Ser264 and D1-His252. A hydrogen bond is formed between the aromatic hydroxyl oxygen of GT and the residue Ser264 in the D1 protein. The 4-carbonyl group of GT provides another hydrogen bond to the residue D1-His252. So, it is concluded that GT is a novel natural PSII inhibitor. In the future, GT may have the potential for development into a bioherbicide or being utilized as a lead compound to design more new derivatives.
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