A complete structural characterization of the desferrioxamine E biosynthetic pathway from the fire blight pathogen Erwinia amylovora

Salomone‐Stagni, Marco; Bartho, J.D.; Polsinelli, Ivan; Bellini, Dom; Walsh, Martin A.; Demitri, Nicola; Benini, Stefano

doi:10.1016/j.jsb.2018.02.002

Cited by 31 publications

(41 citation statements)

References 75 publications

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“…The authors of ar ecently published crystal structure of the desferrioxamine synthetase DfoC (sharing only 54 %s equence identity with DesD) concluded that smalls cale changes of amino acid side chains rather than large structural variations would be responsible for differences in reactions of these type CN IS synthetases. [26] Our results also implicate ar apid route for the evolution and diversificationo fs iderophores. We have demonstrated that only as mall number of mutationsc an significantly alter the product spectrum and that this is in accordance with our model in which the affinity of two adjacent binding sites deter-mines oligomerization and macrocyclization steps.…”

Section: Discussionsupporting

confidence: 60%

“…We suspected that minor changes of amino acids in the mutant modulated the affinity of the substrate binding sites. The authors of a recently published crystal structure of the desferrioxamine synthetase DfoC (sharing only 54 % sequence identity with DesD) concluded that small scale changes of amino acid side chains rather than large structural variations would be responsible for differences in reactions of these type C NIS synthetases …”

Section: Resultsmentioning

confidence: 99%

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Dissecting the Mechanism of Oligomerization and Macrocyclization Reactions of NRPS‐Independent Siderophore Synthetases

Rütschlin

Böttcher

2018

Chemistry A European J

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Macrocyclic and linear hydroxamate siderophores produced by NRPS-independent siderophore (NIS; NRPS=nonribosomal peptide synthetase) synthetases are important in the bacterial competition for iron, as virulence factors, and as drugs for medical use in humans. Despite their importance, the mechanistic details of NIS synthetases have so far remained obscure. Using synthetic substrate analogues as tools allowed for an interrogation of the mechanism of the two closely related NIS synthetases AvbD and DesD. While AvbD produces macrocyclic homo- and heterodimers as native products, DesD is responsible for the synthesis of trimeric desferrioxamines. These enzymes comprise two adjacent binding sites with different substrate selectivities, which direct oligomerization and macrocyclization steps. Exploiting this difference, synthetic substrates were used to invert the native affinities for the sites resulting in switching from trimerization to dimerization reactions for DesD. Based on this work, a comprehensive model explaining the mechanistic details of the reactions and the differences between trimerizing and dimerizing enzymes was developed. Finally, a DesD mutant demonstrated the tuneability of the enzyme's substrate selectivity by only minor changes in the protein sequence. This finding confirms the affinity-directed mechanism responsible for the iterativity of oligomerization and macrocyclization steps.

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Section: Discussionsupporting

confidence: 60%

Section: Resultsmentioning

confidence: 99%

Dissecting the Mechanism of Oligomerization and Macrocyclization Reactions of NRPS‐Independent Siderophore Synthetases

Rütschlin

Böttcher

2018

Chemistry A European J

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“…Cadaverine is immediately monooxygenated by DfoA, thus generating N-hydroxyl-cadaverine. In the last steps DfoC first catalyzes the condensation of N-hydroxyl cadaverine with a succinyl moiety, to give N-5-aminopentyl-N-(hydroxyl)succinamic acid, which is then promptly processed by DfoC synthetase domain, for trimerization and cyclization, to give desferrioxamine E. Beside the structural characterization, in vitro activity of DfoA and DfoJ was also tested in the same work, to confirm the role of these enzymes in the pathway (Salomone-Stagni et al 2018b).…”

Section: Desferrioxamine Pathway Dfoj Dfoa Dfocmentioning

confidence: 99%

“…DfoA Asp392 sidechain is in a perfect position to bind cadaverine N7 amino group. Furthermore, Thr241(Oγ) may form a hydrogen bond with cadaverine N7 amino group (Salomone-Stagni et al 2018b).…”

Section: Desferrioxamine Pathway Dfoj Dfoa Dfocmentioning

confidence: 99%

“…Access to the NTP binding site would be blocked when the N-5-aminopentyl-N-(hydroxyl)-succinamic acid substrate is loaded. The authors identified a "channel" that would allow NTP recycling without requiring the major substrates to be released (Salomone-Stagni et al 2018b). DfoC structure features a large cavity bridging across the DfoC dimerization interface that could allow the 'head' of N-5-aminopentyl-N-(hydroxyl)succinamic acid to enter into the binding pocket of one subunit, and the 'tail' to enter into the binding pocket of the other subunit, therefore suggesting a cooperative synthesis of the DFO-E ring structure (Salomone-Stagni et al 2018a, b).…”

Section: Desferrioxamine Pathway Dfoj Dfoa Dfocmentioning

confidence: 99%

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Structural and functional characterization of proteins from the fire blight pathogen Erwinia amylovora. A review on the state of the art

Benini

2020

J Plant Pathol

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Together with genome analysis and knock-out mutants, structural and functional characterization of proteins provide valuable hints on the biology of the organism under investigation. Structural characterization can be achieved by techniques such as X-ray crystallography, NMR, Cryo-EM. The information derived from the structure are a good starting point to comprehend the details of the proteins molecular function for a better understanding of their biological role. This review aims at describing the progress in the structural and functional characterization of proteins from the plant pathogen Erwinia amylovora obtained by structural biology and currently deposited in the Protein Data Bank.

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Flavin-dependent N-hydroxylating enzymes: distribution and application

Mügge

Heine

Baraibar

et al. 2020

Appl Microbiol Biotechnol

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Amino groups derived from naturally abundant amino acids or (di)amines can be used as "shuttles" in nature for oxygen transfer to provide intermediates or products comprising NO functional groups such as N-hydroxy, oxazine, isoxazolidine, nitro, nitrone, oxime, C-, S-, or N-nitroso, and azoxy units. To this end, molecular oxygen is activated by flavin, heme, or metal cofactorcontaining enzymes and transferred to initially obtain N-hydroxy compounds, which can be further functionalized. In this review, we focus on flavin-dependent N-hydroxylating enzymes, which play a major role in the production of secondary metabolites, such as siderophores or antimicrobial agents. Flavoprotein monooxygenases of higher organisms (among others, in humans) can interact with nitrogen-bearing secondary metabolites or are relevant with respect to detoxification metabolism and are thus of importance to understand potential medical applications. Many enzymes that catalyze N-hydroxylation reactions have specific substrate scopes and others are rather relaxed. The subsequent conversion towards various NO or N-N comprising molecules is also described. Overall, flavin-dependent N-hydroxylating enzymes can accept amines, diamines, amino acids, amino sugars, and amino aromatic compounds and thus provide access to versatile families of compounds containing the NO motif. Natural roles as well as synthetic applications are highlighted. Key points • NO and N-N comprising natural and (semi)synthetic products are highlighted. • Flavin-based NMOs with respect to mechanism, structure, and phylogeny are reviewed. • Applications in natural product formation and synthetic approaches are provided.

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A complete structural characterization of the desferrioxamine E biosynthetic pathway from the fire blight pathogen Erwinia amylovora

Cited by 31 publications

References 75 publications

Dissecting the Mechanism of Oligomerization and Macrocyclization Reactions of NRPS‐Independent Siderophore Synthetases

Dissecting the Mechanism of Oligomerization and Macrocyclization Reactions of NRPS‐Independent Siderophore Synthetases

Structural and functional characterization of proteins from the fire blight pathogen Erwinia amylovora. A review on the state of the art

Flavin-dependent N-hydroxylating enzymes: distribution and application

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