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Chapter 60 : Genetic and Structural Examination of the Legiobactin Siderophore
Bacteria have evolved numerous ways to acquire iron from the environment, but a common mechanism is the production of siderophores. Siderophores are electronegative, iron-regulated, low-molecular-weight compounds produced by bacteria and fungi that bind ferric iron and facilitate its internalization via specific receptors. Siderophores are secondary metabolites mainly synthesized by nonribosomal peptide synthetases that are similar to those used for antibiotic synthesis. Iron-bound siderophores are usually recognized at the cell surface by specific receptors in both gram-positive and gram-negative bacteria that internalize the ferrisiderophore. Siderophores are extremely effective in binding the Fe3+ ion because they contain the most effective iron binding ligands in nature, consisting of hydroxamate, catecholate, and α-hydroxycarboxylate ligands that form hexadentate Fe3+ complexes, thus satisfying the six coordination sites on ferric ions. The authors showed that L. pneumophila could produce a high-affinity iron-chelator. When grown at 37°C in a low-iron chemically defined medium (CDM), L. pneumophila secretes a low-molecular-weight substance that is reactive in the Chrome Azurol S (CAS) assay. Indeed, legiobactin does not extract into common solvents that are used to extract catecholate and hydroxamate siderophores. The legiobactin peak is the portion of L. pneumophila supernatants that promotes growth of iron-starved legionellae and is absent in the lbtA mutant supernatants. As was the case using 13 C nuclear magnetic resonance (NMR) analysis, proton NMR analysis of purified legiobactin demonstrates that the siderophore contains only aliphatic residues. Currently, the authors are working toward determining the structure of legiobactin based on two-dimensional-NMR, elemental analysis, and Maldi experiments.