Nickel ions (Ni(II)) are required cofactors for a number of microbial enzymes. Under anaerobic conditions E. coli requires nickel for its Ni/Fe-hydrogenase enzymes. E. coliobtains Ni(II) from its surroundings with the nickel-specific transporter NikABCDE. However excess nickel is potentially toxic and necessitates tight control of nickel influx. The NikR protein is a nickel- and DNA-binding transcriptional repressor that responds to elevated levels of intracellular Ni(II) by binding to the nikABCDE operator DNA and repressing nickel transporter production. NikR is a homotetramer with 4 Ni(II) -binding sites clustered at the tetrameric interface (~1pM binding affinity), and 2 DNA-binding sites at opposite ends of the molecule. Ni(II)-binding activates NikR binding to nikABCDE operator DNA with ~20nM affinity. Based on the crystal structure of NikR, approximately 40 angstroms separates the DNA- and Ni(II)-binding sites, and the mechanism by which Ni(II) activates the NikR protein for DNA-binding remains unknown. Using a combination of computational and experimental methods we are characterizing the network of amino acid residue interactions necessary for NikR activation. Molecular modeling strategies including atomic-scale molecular dynamics and coarse-grain modeling of large conformational changes are utilized to predict residues important for NikR function and suggest a mechanism of Ni(II)-dependent activation. Hydrogen/deuterium exchange measured by mass spectrometry is being developed to measure NikR conformational change and validate the simulations. Important amino acid residues identified in the simulations will be mutated, and mutant NikR proteins will be tested experimentally for changes in nickel binding, the capacity to undergo Ni(II)-dependent conformational change, and the capability to bind DNA.