The Aer aerotaxis receptor in Escherichia coli mounts such a rapid response to redox that E. coli changes its swimming behavior within 100 ms after an oxygen pulse. This receptor is a membrane-bound homodimer, and it monitors internal energy (redox) via an FAD cofactor bound to a cytoplasmic N-terminal PAS domain. Understanding PAS sensing is important, as PAS domains comprise a superfamily of more than 25,000 members from all kingdoms. This study focused on the Aer N-terminal PAS sensor, as well as the region tethering it to the membrane anchor, known as the F1. Previous genetic studies on these regions identified several residues that are important for structure and function, but the number and type of mutants isolated were too few to propose how signaling might occur within the PAS sensor. In this study, I used random mutagenesis to identify additional residues in the PAS domain that are critical for FAD-binding, sensing and signal transduction. I found that Aer mutant proteins did not require FAD binding for maturation and stability. However, lesions that appeared to disrupt PAS-HAMP interaction and/or proper folding of the FAD binding pocket inhibited maturation and increased proteolysis. The most consequential lesions isolated were gain-of-function mutations, because they mimicked the signal-on state of the receptor, and allowed us to propose a pathway through the PAS sensor itself. When mapped onto a homology model, these 24 mutations localized to the FAD-binding cleft, the β-scaffold and the loop connecting the N-terminal cap to the PAS core. We propose that the reduction of FAD initiates a conformational change in the FAD binding pocket that repositions the β-scaffolding and the N-cap loop. The residues surrounding the β-scaffolding and N-cap loop in turn may be important for transmitting the conformational signal directly to the cytoplasmic HAMP and signaling domain. To investigate the function and structure of the F1 region of the Aer receptor, we again used random mutagenesis, but also employed cysteine scanning, disulfide crosslinking, as well as heterodimers composed of a defective full-length receptor and a truncated monomer retaining different regions of the F1. I found that the proximal end of the F1 forms an N-terminal helix followed by an extended central loop. At this central loop, cognate F1s were close and interacted between subunits. From the lesions isolated in this region we concluded that the F1 region was important for Aer stability rather than function. But experiments with heterodimers also indicated that native F1 residues spanning 154 to 164 were required for function, in addition to the presence of at least one arginine residue at the membrane interface. From these data, as well as ab initio and homology models of the PAS, N-terminal region of the F1 (both based on MmoS) and the HAMP domain (based on Af1503), we constructed a working model of the Aer trimer of dimers. Based on this model, the PAS domains and F1 regions circumscribe the HAMP domain, and the F1 regions interact with neighboring dimers, both within a trimer of dimers, as well as between trimers of dimers.

LLU Discipline

Microbiology and Molecular Genetics


Microbiology, Molecular Biology and Biochemistry


Graduate Studies

First Advisor

Barry L. Taylor

Second Advisor

Katherine A. Borkovich

Third Advisor

Alan Escher

Fourth Advisor

Mark S. Johnson

Fifth Advisor

Subburaman Mohan

Sixth Advisor

Ubaldo Soto

Seventh Advisor

Kylie J. Watts

Degree Name

Doctor of Philosophy (PhD)

Degree Level


Year Degree Awarded


Date (Title Page)




Library of Congress/MESH Subject Headings

Aer protein, E coli; Escherichia coli Proteins -- genetics; Mutant Proteins -- genetics; Carrier Proteins -- genetics; Protein Binding; Receptors, Aryl Hydrocarbon; Signal Transduction; DNA Mutational Analysis; Molecular Sequence Data; Models, Molecular; Models, Biological.



Page Count

xx; 196

Digital Format


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.


Loma Linda University Electronic Theses and Dissertations

Collection Website



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