EPR spectroscopy detects various active state conformations of the copper regulator CueR

The interactions between proteins and their specific DNA sequences are the genesis of many cellular processes. Hence, developing methods designed to build an atomic level picture of these interactions help us improve the understanding of key cellular mechanisms. CueR protein is an Escherichia coli copper sensing transcription regulator. Copper regulators have been the focus of attention for the past few years, since their inhibition can kill pathogenic systems, without harming the host cell. Several spectroscopic tools and crystallographic data suggested that conformational changes induced both by the DNA and the protein control the transcription process. However, due to the inadequate resolution of these methods, the exact number of protein conformations at the active state have not been determined and characterized. Resolving the structure of CueR at the active state is highly important in order to be able to identify a specific site for inhibition. Here, we show the potential of double-histidine (dHis) based Cu(II) spin-labelling to identify various conformational states of CueR during the transcription process.