Engineering the Escherichia coli NikR Transcription Factor via Mutations in the α3 Helix

by Seyedeh Mahshid Mortazavi

Institution: University of Toronto
Year: 2014
Record ID: 2024760
Full text PDF: http://hdl.handle.net/1807/65590


Escherichia coli NikR is a metal-responsive transcription factor that controls nickel uptake by regulating expression of a nickel-specific transporter, NikABCDE. NikR contains a high-affinity nickel-binding site, and nickel must occupy this site in order to activate DNA binding. Stoichiometric amounts of nickel bind to this site and stabilize the α3 helix and the preceding loop (residues 63 – 79). The α3 helix forms non-specific electrostatic interactions with the DNA. The current hypothesis is that nickel activates NikR-DNA binding by organizing α3 helix and activating non-specific DNA contacts, allowing NikR to scan along the DNA until it reaches the nik promoter. This presentation will examine this hypothesis by generating a nickel independent NikR through mutagenesis.