CAREER: Structural and Mechanistic Studies on a Iron-Sulfur Cluster-Based Nitric Oxide Sensor

University Of Nebraska-Lincoln, Lincoln NE

Investigators

Abstract

Because iron (Fe) and sulfur (S) ions are both highly reactive, proteins that contain multi-iron and sulfur assemblies (Fe-S clusters), can play versatile roles in how bacteria sense and rapidly adapt to environmental stress. Proper function of Fe-S cluster proteins is required for the fitness of bacteria. These proteins often confer persistence and antibiotic resistance to bacteria that are pathogenic to their host. Despite the importance of these Fe-S cluster proteins, there are critical gaps in knowledge about how they function in bacteria and their underlying mechanisms of action due to the technical challenges of working with these oxygen-sensitive proteins. The Chemistry of Life Processes and Established Program to Stimulate Competitive Research (EPSCoR) Programs jointly fund Dr. Limei Zhang at the University of Nebraska-Lincoln. Professor Zhang builds an integrated research and education program whose goal is to decipher the structural basis for the unique mode of action of a very important, but understudied, Fe-S cluster protein in Actinobacteria called WhiB1. The knowledge gained from this project may help combat pathogenic actinobacteria such as the deadly, persistent pathogen Mycobacterium tuberculosis. The research may also improve production of bioactive molecules using actinobacteria that could, in turn, have benefits for the agriculture. This research is integrated into a series of education and outreach activities designed to recruit and train next-generation STEM researchers, with an emphasis on groups underrepresented in science. It also promotes science literacy by engaging with audiences that include K-12, undergraduate, and graduate students, as well as the general public. WhiB1 is a [4Fe-4S] cluster-based nitric oxide (NO) sensor widely distributed in actinobacteria and essential in Mycobacterium tuberculosis (Mtb). WhiB1 is linked to the NO stress response and initiation of dormancy in Mtb. Despite decades of investigation on this protein, crucial knowledge gaps remain in defining the mode of action of WhiB1 for its essential role in supporting active cell growth and for selectively sensing and regulating gene expression in response to cellular NO levels under aerobic conditions. Beyond the details about NO sensing and stress response, WhiB1 represents a large family of understudied monomeric transcription factors involved in signaling, of which the underlying mechanisms of action are unknown. In this project, Dr. Zhang and her research group are using Mtb WhiB1 as the model protein and a suite of high-end structural, spectroscopic, and molecular tools to advance mechanistic understanding of the [4Fe-4S] cluster-dependent control of protein folding and dynamics; the NO-selectivity of the [4Fe-4S] cluster-based transcription factors; and the mode of action of the WhiB1-like monomeric transcription factors. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.