Allosteric Regulation of the Nickel-dependent NikR Repressor

Washington University School Of Medicine, Saint Louis MO

Investigators

Abstract

Intellectual merit. Transition metals are an essential nutrient for all living cells, yet they are toxic in excess amounts. Organisms utilize several mechanisms to ensure metal levels are carefully controlled. Microbes commonly use metal-sensing DNA-binding proteins to control transcriptional regulation of metal transporters, thereby regulating the amount of metal transport in response to intracellular metal levels. This project focuses on the nickel-dependent NikR transcriptional regulator, a protein found in a wide variety of bacterial and archaeal species. The protein contains two distinct structural modules: an N-terminal DNA-binding domain and a C-terminal nickel binding domain. NikR binds DNA only in the presence of nickel and thus serves to sense excess nickel inside the cell. This project seeks to understand how nickel alters the NikR structure to enable DNA binding. Using a combination of computational and experimental approaches, the amino acid residues required for allosteric communication between the nickel- and DNA-binding domains will be identified. Computational methods will characterize interaction networks and large scale motions that are altered in the presence of nickel. Mutagenesis approaches will be used to identify amino acids that are critical for the nickel-dependent response of NikR. The experiments are designed to either disrupt these interactions or, using the power of genetic approaches, identify mutants that bypass the nickel requirement for DNA-binding. Structural mapping of these residues provides a point of contact with computational methods. Broader impacts. The interdisciplinary nature of this project has a direct impact on the training of students by providing access to a wider variety of experimental methodologies. The experimental objectives of the project are well suited to early research experiences by exposing to fundamental concepts in microbial physiology and genetics, protein structure and function, and bioinorganic chemistry. Participants in this research include students from the Students and Teachers as Research Scientists (STARS) summer program and the Washington University Young Scientist Program (YSP) both of which provide summer research experiences for St. Louis area high school students from diverse backgrounds.