Cytoplasmic sensing by Bacillus subtilis chemoreceptors

Motile bacteria navigate chemical cues in their surroundings using a process called chemotaxis. They employ a modified two component system to sense and respond to chemical gradients in the environment. This system comprises of chemoreceptors that detect chemicals or ligands. Chemoreceptors are generally transmembrane proteins that have evolved to form diverse extracellular ligand binding structures. The canonical mode of sensing is via direct binding of ligands to these ligand binding domains to induce conformational changes that are propagated to the flagellar motor. This dissertation will present my work on identifying an unconventional sensing mechanism in which ligands bind to the conserved cytoplasmic domain of the chemoreceptor instead of the extracellular ligand binding domain. Chapter 1 gives a brief introduction on the different components of the chemotaxis pathway with an emphasis on the gram-positive model organism, Bacillus subtilis. This chapter also discusses different sensing mechanisms reported in the literature. Chapter 2 is a report on the cytoplasmic sensing mechanism involved in chemotaxis towards ethanol by B. subtilis. Using chimeric receptors, we showed that ethanol interacts with the conserved signaling domain of McpB chemoreceptor. This was corroborated in vitro using Saturation Transfer Difference NMR experiments. Molecular Dynamics simulations performed by Dr. C. Keith Cassidy, showed the importance of a single reside, Ala431, involved in sensing ethanol. Chapter 3 outlines my work on repellent chemotaxis in B. subtilis. Using capillary assay experiments, we found that phenol and related aromatics are sensed as attractants at low concentrations and repellents at high concentrations. McpA chemoreceptor was found to mediate repellent chemotaxis response to a variety of aromatic compounds. Chimeric receptor analysis and in vitro protein ligand binding experiments showed that the cytoplasmic region of McpA chemoreceptor is involved in sensing these molecules. Chapter 4 provides a summary of preliminary results on chemotaxis towards plastic leachates by B. subtilis. Polyethylene terephthalate bottles leach certain chemicals that are sensed by B. subtilis as attractants. The response is variable and dependent on the brand of water bottle, and the method of leachate preparation. Chapter 5 is an outline of my attempt to design a chemoreceptor to sense a non-native ligand using the Rosetta modelling suite. Screening assays are discussed with an outline of future directions for this project. Chapter 6 presents the major conclusions and an outlook on the bacterial chemotaxis field.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Chemical Engineering

Grantor

University of Illinois at Urbana-Champaign

Year dc:date

2022

Author dc:creator

• Bodhankar, Girija

Contributors dc:contributor

• Rao, Christopher
• Kraft, Mary
• Harley, Brendan
• Chemla, Yann

• Chemotaxis
• Protein ligand binding
• Bacillus subtilis
• STD NMR

Statement dc:rights

• © 2022 by Girija Bodhankar. All rights reserved.

Language dc:language

en, eng

Handle dc:identifier

https://hdl.handle.net/2142/115492

OAI record oai:identifier

https://www.ideals.illinois.edu/oai-pmh?verb=GetRecord&metadataPrefix=etdms&identifier=oai%3Awww.ideals.illinois.edu%3A2142%2F115492