Ferri-FSC (Fusarine C, Fusigen) is a siderophore made of three N5-anhydromevalonyl-N5-hydroxyornithine residues, termed fusarinine (FS), which is cyclically linked by ester bonds. Both extra- and intracellular siderophores have been shown to be crucial for the virulence of A. fumigatus. Subsequent to chelation of iron and uptake, FSC is hydrolyzed and the iron is transferred to the metabolism or to the intracellular siderophore FC for transport and storage of iron. SidJ is an esterase which hydrolyse Ferri-FSC. However this protein is only representative of trypanosomatid members of this sequence family as these proteins lack some if not all of the catalytic triad residues. TafC esterase EstB of from Aspergillus fumigatus does not hydrolyses fusigen-Fusarinine C
Fusarinine C (FSC), a siderophore-based chelator coupled with the model peptide c(RGDfK) (FSC(succ-RGD)3), revealed excellent targeting properties in vivo using positron emission tomography (PET). Here, we report the details of radiolabeling conditions and specific activity as well as selectivity for (68)Ga. (68)Ga labeling of FSC(succ-RGD)3 was optimized regarding peptide concentration, pH, temperature, reaction time, and buffer system. Specific activity (SA) of [(68)Ga]FSC(succ-RGD)3 was compared with (68)Ga-1,4,7-triazacyclononane, 1-glutaric acid-4,7 acetic acid RGD ([(68)Ga]NODAGA-RGD). Stability was evaluated in 1000-fold ethylenediaminetetraacetic acid (EDTA) solution (pH 7) and phosphate-buffered saline (PBS). Metal competition tests (Fe, Cu, Zn, Al, and Ni) were carried out using [(68)Ga]-triacetylfusarinine C. High radiochemical yield was achieved within 5 min at room temperature, in particular allowing labeling with (68)Ga up to pH 8 with excellent stability in 1000-fold EDTA solution and PBS. The 10-fold to 20-fold lower concentrations of FSC(succ-RGD)3 led to the same radiochemical yield compared with [(68)Ga]NODAGA-RGD with SA up to 1.8 TBq/micromol. Metal competition tests showed high selective binding of (68)Ga to FSC. FSC is a multivalent siderophore-based bifunctional chelator allowing fast and highly selective labeling with (68)Ga in a wide pH range and results in stable complexes with high SA. Thus it is exceptionally well suited for the development of new (68)Ga-tracers for in vivo molecular imaging with PET.
Title: Fungal siderophore metabolism with a focus on Aspergillus fumigatus Haas H Ref: Nat Prod Rep, 31:1266, 2014 : PubMed
Siderophores are chelators synthesized by microbes to sequester iron. This article summarizes the knowledge on the fungal siderophore metabolism with a focus on Aspergillus fumigatus. In recent years, A. fumigatus became a role model for fungal biosynthesis, uptake and degradation of siderophores as well as regulation of siderophore-mediated iron handling and the elucidation of siderophore functions. Siderophore functions comprise uptake, intracellular transport and storage of iron. This proved to be crucial not only for adaptation to iron starvation conditions but also for germination, asexual and sexual propagation, antioxidative defense, mutual interaction, microbial competition as well as virulence in plant and animal hosts. Recent studies also indicate the high potential of siderophores and its biosynthetic pathway to improve diagnosis and therapy of fungal infections.
Siderophore-mediated iron handling is crucial for the virulence of Aspergillus fumigatus. Here we identified a new component of its siderophore metabolism, termed SidJ, which is encoded by AFUA_3G03390. The encoding gene is localized in a siderophore biosynthetic gene cluster that is conserved in a variety of fungi. During iron starvation, SidJ deficiency resulted in decreased growth and increased intracellular accumulation of hydrolysis products of the siderophore fusarinine C. The implied role in siderophore hydrolysis is consistent with a putative esterase domain in SidJ, which now represents the first functionally characterized member of the DUF1749 (domain of unknown function) protein family, with members found exclusively in fungi and plants.
1 lessTitle: Iron Scavenging in Aspergillus Species: Structural and Biochemical Insights into Fungal Siderophore Esterases Ecker F, Haas H, Groll M, Huber EM Ref: Angew Chem Int Ed Engl, 57:14624, 2018 : PubMed
Fungi utilize high-affinity chelators termed siderophores with chemically diverse structures to scavenge the essential nutrient iron from their surroundings. Since they are among the strongest known Fe(3+) binding agents, intracellular release of the heavy metal atom is facilitated by the activity of specific hydrolases. In this work, we report the characterization and X-ray crystal structures of four siderophore esterases: AfEstB and AfSidJ from Aspergillus fumigatus, as well as AnEstB and AnEstA from Aspergillus nidulans. Even though they all display the conserved alpha/beta-hydrolase fold, we found significant structural and enzymatic discrepancies in their adaption to both related and chemically diverse substrates. A structure of AfEstB in complex with its substrate triacetylfusarinine C gives insight into the active enzyme and shows tetrahedral coordination between the catalytic serine and the scissile ester bond.
Fusarinine C (FSC), a siderophore-based chelator coupled with the model peptide c(RGDfK) (FSC(succ-RGD)3), revealed excellent targeting properties in vivo using positron emission tomography (PET). Here, we report the details of radiolabeling conditions and specific activity as well as selectivity for (68)Ga. (68)Ga labeling of FSC(succ-RGD)3 was optimized regarding peptide concentration, pH, temperature, reaction time, and buffer system. Specific activity (SA) of [(68)Ga]FSC(succ-RGD)3 was compared with (68)Ga-1,4,7-triazacyclononane, 1-glutaric acid-4,7 acetic acid RGD ([(68)Ga]NODAGA-RGD). Stability was evaluated in 1000-fold ethylenediaminetetraacetic acid (EDTA) solution (pH 7) and phosphate-buffered saline (PBS). Metal competition tests (Fe, Cu, Zn, Al, and Ni) were carried out using [(68)Ga]-triacetylfusarinine C. High radiochemical yield was achieved within 5 min at room temperature, in particular allowing labeling with (68)Ga up to pH 8 with excellent stability in 1000-fold EDTA solution and PBS. The 10-fold to 20-fold lower concentrations of FSC(succ-RGD)3 led to the same radiochemical yield compared with [(68)Ga]NODAGA-RGD with SA up to 1.8 TBq/micromol. Metal competition tests showed high selective binding of (68)Ga to FSC. FSC is a multivalent siderophore-based bifunctional chelator allowing fast and highly selective labeling with (68)Ga in a wide pH range and results in stable complexes with high SA. Thus it is exceptionally well suited for the development of new (68)Ga-tracers for in vivo molecular imaging with PET.
Title: Fungal siderophore metabolism with a focus on Aspergillus fumigatus Haas H Ref: Nat Prod Rep, 31:1266, 2014 : PubMed
Siderophores are chelators synthesized by microbes to sequester iron. This article summarizes the knowledge on the fungal siderophore metabolism with a focus on Aspergillus fumigatus. In recent years, A. fumigatus became a role model for fungal biosynthesis, uptake and degradation of siderophores as well as regulation of siderophore-mediated iron handling and the elucidation of siderophore functions. Siderophore functions comprise uptake, intracellular transport and storage of iron. This proved to be crucial not only for adaptation to iron starvation conditions but also for germination, asexual and sexual propagation, antioxidative defense, mutual interaction, microbial competition as well as virulence in plant and animal hosts. Recent studies also indicate the high potential of siderophores and its biosynthetic pathway to improve diagnosis and therapy of fungal infections.
Siderophore-mediated iron handling is crucial for the virulence of Aspergillus fumigatus. Here we identified a new component of its siderophore metabolism, termed SidJ, which is encoded by AFUA_3G03390. The encoding gene is localized in a siderophore biosynthetic gene cluster that is conserved in a variety of fungi. During iron starvation, SidJ deficiency resulted in decreased growth and increased intracellular accumulation of hydrolysis products of the siderophore fusarinine C. The implied role in siderophore hydrolysis is consistent with a putative esterase domain in SidJ, which now represents the first functionally characterized member of the DUF1749 (domain of unknown function) protein family, with members found exclusively in fungi and plants.
