סמינר בכימיה פיזיקלית: The use of EPR spectroscopy to study metal transcription factors in-vitro and in-cell

​Abstract:

Bacteria are skilled survivors in high-metal ion environments. Some metals are essential but can be detrimental above a certain concentration in the cytosol. Thus, bacteria have evolved complex processes to regulate intracellular metal homeostasis.  Metal transcription factors play a role in this regulation. They respond to metal ions and control gene transcription to protect bacteria from metal-induced stress. Upon metal binding, the metalloregulator protein induces conformational changes in the DNA promoter sequence, leading to transcription initiation and subsequent transcription of genes that mediate adaptive responses to the toxicity of the targeted metal. X-ray crystallography and cryo-EM tools have provided insights into the structures of these transcription factors bound and unbound to the DNA and metal ions. However, intermediate conformations, which offer a better understanding on their mechanism of action, have not been targeted by these methods. Electron paramagnetic resonance (EPR) spectroscopy can help bridge these gaps. Our lab focuses on studying three copper-sensitive transcription factors from gram-positive and gram-negative bacteria: E. coli and P. aeruginosa CueR 1-8, M. tuberculosis CsoR 9, and S. pneumoniae CopY 10. Using variety of spin-labelling techniques and EPR measurements on both protein and DNA, both in-vitro and in-cell, we demonstrate how metal ions initiate conformational changes that regulate gene transcription. 

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מארגן הסמינר: ד"ר גיא כהן