Erwin Stuffle

Abstract

Bacteria use chemoreceptors and associated chemosensory systems to navigate and respond to their environment. Aer2 chemoreceptors are a family of O2 sensors that all have a common core structure but differ with regards to their N-terminal domain types and arrangements. The Aer2 receptors from Pseudomonas aeruginosa (PaAer2) and Vibrio cholerae (VcAer2) have been extensively studied. However, the marine pathogen Vibrio vulnificus possesses a previously uncharacterized Aer2 receptor. Here I present an analysis of the V. vulnificus Aer2 chemoreceptor, VvAer2, which is the first V. vulnificus chemoreceptor to be characterized. VvAer2 is related to the Aer2 receptors of other gammaproteobacteria, but uncharacteristically contains three PAS domains (PAS1-3), rather than one or two PAS domains. PAS domains are sensing, signaling, dimerization, and protein interaction domains found in proteins from all five kingdoms of life. Using an E. coli chemotaxis hijack assay, I determined that VvAer2, like other Aer2 receptors, senses and responds to O2. All three VvAer2 PAS domains bound pentacoordinate b-type heme and exhibited similar O2 affinities. PAS2 and PAS3 both stabilized O2 via conserved Ib-Trp residues, but PAS1, which was easily oxidized in vitro, was unaffected by Ib-Trp replacement. Collectively, my results support a model in which PAS1 is largely dispensable for O2-mediated signaling, whereas PAS2 modulates PAS3 signaling, and PAS3 signals to the downstream domains. Each PAS domain appears to be positionally optimized, since PAS swapping caused altered signaling properties, and neither PAS1 nor PAS2 could replace PAS3.

I also explored the conservation of signaling mechanisms across Aer2 receptors. I introduced and tested residue replacements in VvAer2 and VcAer2 that were previously characterized in PaAer2, and I created and tested chimeric Vibrio Aer2 receptors. Overall, I determined that Aer2 signaling involves a Cterminal PAS-heme domain that binds O2 and signals via largely conserved mechanisms. However, each Aer2 receptor has evolved to work efficiently with its own complement of N-terminal domains and arrangement. These distinct Nterminal domain arrangements facilitate, modulate and tune responses based on environmental signals.

LLU Discipline

Microbiology

Department

Basic Sciences

School

School of Medicine

First Advisor

Mark S. Johnson

Second Advisor

Kylie J. Watts

Third Advisor

Paul C. Herrmann

Fourth Advisor

Suzanne E. Phillips

Fifth Advisor

Daniel K. Rogstad

Sixth Advisor

Elaine A. Vanterpool

Degree Name

Doctor of Philosophy (PhD)

Degree Level

Ph.D.

Year Degree Awarded

2023

Date (Title Page)

1-2023

Language

English

Library of Congress/MESH Subject Headings

Vibrio vulnificus - chemistry; Chemoreceptor Cells

Type

Dissertation

Page Count

xvii, 138 p.

Digital Format

PDF

Digital Publisher

Loma Linda University Libraries

Usage Rights

This title appears here courtesy of the author, who has granted Loma Linda University a limited, non-exclusive right to make this publication available to the public. The author retains all other copyrights.

Collection

Loma Linda University Electronic Theses and Dissertations

Collection Website

http://scholarsrepository.llu.edu/etd/

Repository

Loma Linda University. Del E. Webb Memorial Library. University Archives

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