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Physical Basis of Metal-Binding Specificity in Escherichia coli NikR

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journal contribution
posted on 2009-07-29, 00:00 authored by Christine M. Phillips, Paul S. Nerenberg, Catherine L. Drennan, Collin M. Stultz
In Escherichia coli and other bacteria, nickel uptake is regulated by the transcription factor NikR. Nickel binding at high-affinity sites in E. coli NikR (EcNikR) facilitates EcNikR binding to the nik operon, where it then suppresses transcription of genes encoding the nickel uptake transporter, NikABCDE. A structure of the EcNikR-DNA complex suggests that a second metal-binding site is also present when NikR binds to the nik operon. Moreover, this co-crystal structure raises the question of what metal occupies the second site under physiological conditions: K+, which is present in the crystal structure, or Ni2+, which has been proposed to bind to low- as well as high-affinity sites on EcNikR. To determine which ion is preferred at the second metal-binding site and the physical basis for any preference of one ion over another in both the second metal-binding site and the high-affinity sites, we conducted a series of detailed molecular simulations on the EcNikR structure. Simulations that place Ni2+ at high-affinity sites lead to stable trajectories with realistic ion−ligand distances and geometries, while simulations that place K+ at these sites lead to conformational changes in the protein that are likely unfavorable for ion binding. By contrast, simulations on the second metal site in the EcNikR-DNA complex lead to stable trajectories with realistic geometries regardless of whether K+ or Ni2+ occupies this site. Electrostatic binding free energy calculations, however, suggest that EcNikR binding to DNA is more favorable when the second metal-binding site contains K+. An analysis of the energetic contributions to the electrostatic binding free energy suggests that, while the interaction between EcNikR and DNA is more favorable when the second site contains Ni2+, the large desolvation penalty associated with moving Ni2+ from solution to the relatively buried second site offsets this favorable interaction term. Additional free energy simulations that account for both electrostatic and non-electrostatic effects argue that EcNikR binding to DNA is most favorable when the second site contains a monovalent ion the size of K+. Taken together, these data suggest that the EcNikR structure is most stable when Ni2+ occupies high-affinity sites and that EcNikR binding to DNA is more favorable when the second site contains K+.

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