Biochemical characterization of ferric uptake regulator (Fur) from Aliivibrio salmonicida . Mapping the DNA sequence spe
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Biochemical characterization of ferric uptake regulator (Fur) from Aliivibrio salmonicida. Mapping the DNA sequence specificity through binding studies and structural modelling Kristel Berg
. Hege Lynum Pedersen
. Ingar Leiros
Received: 16 November 2019 / Accepted: 28 June 2020 Ó The Author(s) 2020
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
Electronic supplementary material The online version of this article (https://doi.org/10.1007/s10534-020-00240-6) contains supplementary material, which is available to authorized users. K. Berg H. L. Pedersen I. Leiros (&) Department of Chemistry, Faculty of Science and Technology, The Norwegian Structural Biology Centre (NorStruct), UiT the Arctic University of Norway, 9037 Tromsø, Norway e-mail: [email protected]
interpreted in the context of homology models of AsFur to gain molecular-level insight into binding specificity. Keywords Ferric uptake regulator Metal binding DNA interactions Aliivibrio salmonicida Electrophoretic mobility shift assay
Introduction Iron is an essential nutrient for all living organisms and many key biological processes are dependent on its abundance. For bacteria, iron is crucial for growth and host colonization. Iron mostly exists in the insoluble Fe3? form under aerobic conditions at physiological pH and availability of the soluble reduced form, Fe2?, is restricted. Due to the ability of free iron to form toxic hydroxyl radicals through the Fenton reaction (Guerinot 1994), the essential high-affinity uptake systems of iron and iron homeostasis in bacteria must be tightly regulated, and in most bacteria, these processes are under control of the global metalloregulator Ferric uptake regulator (Fur) (Hantke 2001). Fur was first described in Escherichia coli (Hantke 1981), where it controls the expression of more than 90 genes, and its chemical properties and role in homeostasis has since been studied in homologs from multiple bacteria, including Mycobacterium smegmatis (Gao et al.
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Biometals
2019), Acidovorax citrulli (Liu et al. 2019), Campylobacter
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