Biochemical characterization of ferric uptake regulator (Fur) from Aliivibrio salmonicida. Mapping the DNA sequence specificity through binding studies and structural modelling

Abstract

Iron is an essential nutrient for bacteria, however its propensity to form toxic hydroxyl radicals at high intracellular concentrations, requires its acquisition to be tightly regulated. Ferric uptake regulator (Fur) is a metal-dependent DNA-binding protein that acts as a transcriptional regulator in maintaining iron metabolism in bacteria and is a highly interesting target in the design of new antibacterial drugs. Fur mutants have been shown to exhibit decreased virulence in infection models. The protein interacts specifically with DNA at binding sites designated as ‘Fur boxes’. In the present study, we have investigated the interaction between Fur from the fish pathogen Aliivibrio salmonicida (AsFur) and its target DNA using a combination of biochemical and in silico methods. A series of target DNA oligomers were designed based on analyses of Fur boxes from other species, and affinities assessed using electrophoretic mobility shift assay. Binding strengths were interpreted in the context of homology models of AsFur to gain molecular-level insight into binding specificity.

Ferric uptake regulator (Fur) binds a [2Fe-2S] cluster to regulate intracellular iron homeostasis in Escherichia coli

“…Binding of Fur to the Fur-box protects the hin fI site from being cleaved by Hin fI ( 18 ). The restriction site protection assay has been applied to investigate the Fur-box binding activity of E. coli Fur after nitric oxide exposure ( 28 ), of the Co(II)-bound Helicobacter pylori Fur ( 21 ), and of the Mn(II)-bound Aliivibrio salmonicida Fur ( 29 ). For the experiments, the promoter sequence of the operon iucABCD was synthesized (GenScript co) and inserted into plasmid pUC19 to create pUC19- iuc ( Fig.…”

Proteomic analysis to unravel the biochemical mechanisms triggered by Bacillus toyonensis SFC 500-1E under chromium(VI) and phenol stress

“…Interestingly, the only transcriptional regulator up-expressed under Cr(VI) was Fur, which is involved in iron homeostasis in response to changes in intracellular levels of Fe 2+ , this last is an indirect chemical Cr(VI) reductants under aerobic environmental conditions(Troxell and Hassan 2013;Bansal et al 2019;Bellenberg et al 2019). Noticeably, Fur was originally described as a repressor of genes involved in iron uptake, storage and metabolism(Berg et al 2020). Recent studies however, have indicated that Fur may also function as an activator(Butcher et al 2012).…”