Structure-Function Relationships in the Gas-Sensing Heme-Dependent Transcription Factors RcoM and DNR

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Autor/in	Bowman, Hannah E.
Titel	Structure-Function Relationships in the Gas-Sensing Heme-Dependent Transcription Factors RcoM and DNR
Quelle	(2016), (293 Seiten) PDF als Volltext Verfügbarkeit Ph.D. Dissertation, University of Wisconsin, Madison
Sprache	englisch
Dokumenttyp	gedruckt; online; Monographie
ISBN	978-1-3399-2354-3
Schlagwörter	Hochschulschrift; Dissertation; Cytology; Biochemistry; Genetics; Molecular Structure; Hazardous Materials; Biology; Metabolism; Diseases; Patients; Science Experiments; Feedback (Response); Prompting; Teaching Methods; Science Instruction + Suchen Sie Ihr Suchwort? Thesis; Dissertations; Academic thesis; Zytologie; Biochemie; Humangenetik; Hazardous substance; Gefahrstoff; Biologie; Stoffwechsel; Disease; Krankheit; Patient; Benutzerführung; Teaching method; Lehrmethode; Unterrichtsmethode; Teaching of science; Science education; Natural sciences Lessons; Naturwissenschaftlicher Unterricht
Abstract	Transition metals play an important role in many biological processes, however, they are also toxic at high concentrations. Therefore, the uptake and efflux of these metals must be tightly regulated by the cell. Bacteria have evolved a variety of pathways and regulatory systems to monitor the presence and concentration of metals in the cellular environment. A key component of those systems are transcription factors that either "sense metals" or use "metal sensors". The first class of these proteins have metals as their allosteric effector ligand. The second class of these proteins utilize transition metal containing cofactors to sense other environmental cues through the specific chemistry afforded by the cofactor. Chapter 1 reviews the current literature regarding both types of transcription factors. The focus of this work has been on two heme-containing, gas-sensing transcription factors found in bacteria, RcoM (regulator of CO metabolism) and DNR (dissimilative nitrate respiration regulator). RcoM is a CO-dependent protein found in "Burkholderia xenovorans" and sits upstream of the "cox" operon for oxidative CO metabolism. RcoM senses the presence of CO, as well as changes in redox potential, through a ligand switch process at its heme cofactor. Chapter 2 details spectroscopic characterization of several methionine mutants to identify the Fe(II) ligand trans to His[superscript 74]. That study concludes that Met[superscript 104] acts as the CO-replacable ligand. Met[superscript 105], while not the ligand, does play an important role in reversibility of the ligand switch process. RcoM has a unique tertiary structure that combines a sensory domain and a DNA-binding domain normally found in two-component systems. Chapter 3 provides evidence that RcoM adopts a dimeric state. Further biophysical and structural characterization gives further insight into how the two domains are organized and the implications for the DNA-binding mechanism. DNR is a NO-sensing transcription factor from "Pseudomonas aeruginosa" and regulates part of the nitrate respiration pathway. "P. aeruginosa" uses this pathway to evade host immune response, especially in cystic fibrosis patients. Previous work makes a strong case for the presence of a heme cofactor in DNR; however, attempts to express large quantities of holo-protein have proven unsuccessful. Chapter 4 outlines strategies used to develop a suitable expression protocol. While not entirely successful, the experiments provide a firm foundation for future research on this protein. During the 2015-2016 school year, I conducted educational psychology research as a project assistant in Prof. Martina Rau's lab. She focuses on understanding how students use visual representations and technology to learn chemistry. Chapter 5 summarizes a study we conducted in CHEM 109 to test whether having a computer provide immediate feedback on wedge-dash drawings as well as prompts to collaborate in lab would lead to learning gains. The effect of the intervention was subtle, but statistically significant. [The dissertation citations contained here are published with the permission of ProQuest LLC. Further reproduction is prohibited without permission. Copies of dissertations may be obtained by Telephone (800) 1-800-521-0600. Web page: http://www.proquest.com/en-US/products/dissertations/individuals.shtml.] (As Provided).
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Update	2020/1/01