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Figure 2
The ability of FhuE to utilize hydroxamate siderophores correlates with in vitro binding affinity. (a) A solid agar growth assay of the E. coli strain lacking other high-affinity uptake systems (E. coli ΔTBDT; see Section 2[link]), illustrating FhuE-dependent growth using either coprogen, rhodotorulic acid or ferrioxamine B as an exclusive source of iron. Plates were spotted with the siderophore solution and the growth of bacteria expressing FhuE (+pFhuE) or not (+pBAD24) was assessed after incubation for 72 h at 24°C. (b) Plot of normalized fluorescence quenching of FhuE at 330 nm at increasing concentrations of siderophore. For Fe-coprogen and Fe-rhodotorulic acid, the observed change in fluorescence is attributable to the quenching of tryptophan residues in the FhuE substrate-binding pocket owing to siderophore binding. F0, fluorescence at 0 µM substrate concentration; F, fluorescence at the listed substrate concentration. Error bars are derived from the standard deviation of three independent experiments.

Volume 6| Part 3| May 2019| Pages 401-411
ISSN: 2052-2525