Abstract
The design, synthesis
and biological evaluation of the artificial enterobactin analogue EntKL
and several fluorophore-conjugates thereof are described. EntKL
provides an attachment point for cargos such as fluorophores
or antimicrobial payloads. Corresponding conjugates are recognized by outer
membrane siderophore receptors of Gram-negative pathogens and retain the
natural hydrolyzability of the tris-lactone backbone, known to be key
for uptake into the cytosol. Initial density-functional theory (DFT)
calculations of the free energies of solvation (ΔG(sol)) and relaxed Fe-O force constants of the
corresponding [Fe-EntKL]3- complexes indicated a similar iron binding
constant compared to natural enterobactin (Ent). The synthesis of EntKL
was achieved via an iterative assembly based on a 3-hydroxylysine building block over 14 steps with an overall yield of
3%. A series of growth recovery assays under iron-limiting conditions with Escherichia coli
and Pseudomonas aeruginosa mutant strains that are defective in natural
siderophore synthesis revealed a potent concentration-dependent growth
promoting effect of EntKL similar to natural Ent.
Additionally, four cargo-conjugates differing in molecular size were able to
restore growth of E. coli indicating an uptake into the cytosol. P. aeruginosa
displayed a stronger uptake promiscuity as six different cargo-conjugates
were found to restore growth under iron-limiting conditions. Imaging studies utilizing
BODIPYFL-conjugates, demonstrated the ability of EntKL
to overcome the Gram-negative outer membrane permeability barrier and thus deliver
molecular cargos via the bacterial iron transport machinery of E. coli
and P. aeruginosa.
Content

Supplementary material

Supporting Information EntKL ChemRxiv