Characterization of β-Hydroxyaspartate Siderophores: Structures, Biosyntheses, Coordination Chemistry, and Photoreactivity
- Author(s): Hardy, Clifford Douglas
- Advisor(s): Butler, Alison
- et al.
Nearly all bacteria require iron to function. Bio-available iron is quite limited in many environments, however. To grow and thrive under iron-limited conditions, bacteria have evolved multiple iron-acquisition strategies. One such strategy is the production of siderophores, small molecule Fe(III)-chelating ligands synthesized by bacteria and exported into the environment. Siderophores bind to environmental Fe(III), and the subsequent metal-ligand complex is recognized by receptors on the surface of the bacterium, whereupon the complex is brought back into the cell, and the iron released for utilization by the microbe. This work is focused on investigations into the structures, biosyntheses, coordination chemistry, and photochemistry of siderophores containing the Fe(III)-binding functional group β-hydroxyaspartate.
Reported herein is the characterization of the new siderophore pacifibactin, predicted through a genome mining approach. Pacifibactin is unusual in that it contains four bidentate metal binding sites, including two β-OHAsp functional groups, and Fe(III), Ga(III), and Zr(IV) coordination chemistry of the ligand is discussed.
All β-OHAsp siderophores are photoreactive when complexed to Fe(III), and the photoreactivity of Fe(III)-pacifibactin is reported. Siderophores containing β-OHAsp are biosynthesized by nonribosomal peptide synthetases (NRPSs), large multi-modular enzymes that assemble peptidic natural products. Two distinct strategies for aspartyl β-hydroxylation in NRPS biosynthesis have evolved—one strategy involves β-hydroxylation by discrete hydroxylase enzymes (termed TβHAsp), the other involves hydroxylase domains with NRPS enzymes (termed IβHAsp). A recent phylogenetic analysis of genes encoding these hydroxylase enzymes and NRPSs containing hydroxylase domains uncovered a striking pattern between phylogeny and the stereoselectivity of the hydroxylases. In this work, three known siderophores (delftibactin, pyoverdine GB-1, and histicorrugatin) are stereochemically characterized for the first time, and the stereochemistry of another siderophore (cupriachelin) is revised. This wealth of new characterization supports and strengthens the known association between phylogeny and stereoselectivity, making it a powerful tool for enhancing structural predictions of NRPS natural products.
Lastly, this work details extensive investigations of siderophore photochemistry. The Fe(III) complexes of two β-OHAsp siderophores, pacifibactin and cupriachelin, are known to be photoreactive, and photoproducts of each complex are known. Reported herein is the
structural characterization of several new photoproducts of both Fe(III)-pacifibactin and Fe(III)-cupriachelin, differing significantly from the previously known photoproducts. The relative abundances of photoproducts observed at differing time points of irradiation are also
described, the first example of a time-dependent analysis of Fe(III)-siderophore photoreactivity. Together these results support a mechanistic hypothesis of successive photolytic decarboxylation events driving Fe(III)-siderophore photoreactions.