2026-09-19T23:13:52Zhttps://dspace.mit.edu/server/oai/requestoai:dspace.mit.edu:1721.1/1306152026-06-16T18:53:24Zcom_1721.1_7582com_1721.1_7581col_1721.1_131022
Ronald T. Raines.
Kilgore, Henry R.(Henry Ralph)
Massachusetts Institute of Technology. Department of Chemistry.
Massachusetts Institute of Technology. Department of Chemistry
2021-05-14T16:30:31Z
2021-05-14T16:30:31Z
2020
https://hdl.handle.net/1721.1/130615
1249684883
Thesis: Ph. D., Massachusetts Institute of Technology, Department of Chemistry, September, 2020
Cataloged from the official PDF of thesis. "September 2020."
Includes bibliographical references (pages 385-409).
Protein modifications endow diverse chemical and biological functions. Development of new modifications, control over extant, and the appreciation of their consequences promises to unveil the mechanisms of disease, provide avenues of therapeutic development, and provide insight into intricacies of life. As a result, developing, harnessing, and understanding protein modifications has led to profound direct and indirect consequences for biotechnology, the pharmacopeia and academic research. The express purpose of this thesis was to characterize and investigate the consequences of these modifications. This led to investigations into the stereoelectronic effects endemic to the properties of cysteines and disulfide bonds in proteins and provided a rational basis for the function of thioredoxins, thioredoxin-fold enzymes, and other enzymes that bear highly variable reduction potentials.
Relationships between the ground state geometric properties of disulfide bonds and their photophysical properties has revealed new relationships with potential applications in photoredox chemistry, sulfur-photocatalysis, and protein engineering and design. A strategy based on stereoelectronic effects was used to diversify the reduction potential innate to the cytotoxic activity of epidithiodiketopiperazine natural products, and is leading to experiments that clarify their mechanism of action in cellulo. Detailed analysis revealed a relationship between the thermodynamics of thioldisulfide electrochemical equilibria and the interaction of these motifs with light. Arising from optimization of the hydrolytic stability of fluorogenic probes, a halogen n-->[pi]* interaction in acylated 22,72-halosubstituted fluorosceins was observed and characterized.
Coincident with the investigations into the physicochemical properties of disulfide bonds and other n-->[pi]* interactions, insights into the biological and functional consequences of glycosylation were also investigated. An apparent difference arose in the internalization and release of different dextrans functionalized with fluorogenic probes, suggesting a glycomonomer oxidation-state dependent mechanism for endosomal uptake and release. Further, transformation of proteins with pendant dextrans endowed increased cellular internalization as assessed with proteins that initiate cell death upon internalization. The structure of glycosylated human ribonuclease 1 afforded insight into the origin of their variable catalytic and thermostability. These investigations unveiled a new helix-capping motif, stemming from N-glycosylation at the C terminus of an α-helix.
Finally, the characterization of cellular reactivity and the associated mass transport properties of SNO-OCT reagents, their ability to engage in triple ligations, and their utility for metabolic labeling experiments were also investigated.
by Henry R. Kilgore.
Ph.D.
Ph.D. Massachusetts Institute of Technology, Department of Chemistry
409 pages
eng
Massachusetts Institute of Technology
MIT theses may be protected by copyright. Please reuse MIT thesis content according to the MIT Libraries Permissions Policy, which is available through the URL provided.
http://dspace.mit.edu/handle/1721.1/7582
Chemistry.
Physical consequences of natural and synthetic post-translational modifications
Thesis
application/pdf
2021-05-14T16:30:31Z
Publication
Doctoral
Chem
unknown
unknown
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<Abstract>Protein modifications endow diverse chemical and biological functions. Development of new modifications, control over extant, and the appreciation of their consequences promises to unveil the mechanisms of disease, provide avenues of therapeutic development, and provide insight into intricacies of life. As a result, developing, harnessing, and understanding protein modifications has led to profound direct and indirect consequences for biotechnology, the pharmacopeia and academic research. The express purpose of this thesis was to characterize and investigate the consequences of these modifications. This led to investigations into the stereoelectronic effects endemic to the properties of cysteines and disulfide bonds in proteins and provided a rational basis for the function of thioredoxins, thioredoxin-fold enzymes, and other enzymes that bear highly variable reduction potentials.</Abstract>
<Abstract>Relationships between the ground state geometric properties of disulfide bonds and their photophysical properties has revealed new relationships with potential applications in photoredox chemistry, sulfur-photocatalysis, and protein engineering and design. A strategy based on stereoelectronic effects was used to diversify the reduction potential innate to the cytotoxic activity of epidithiodiketopiperazine natural products, and is leading to experiments that clarify their mechanism of action in cellulo. Detailed analysis revealed a relationship between the thermodynamics of thioldisulfide electrochemical equilibria and the interaction of these motifs with light. Arising from optimization of the hydrolytic stability of fluorogenic probes, a halogen n-->[pi]* interaction in acylated 22,72-halosubstituted fluorosceins was observed and characterized.</Abstract>
<Abstract>Coincident with the investigations into the physicochemical properties of disulfide bonds and other n-->[pi]* interactions, insights into the biological and functional consequences of glycosylation were also investigated. An apparent difference arose in the internalization and release of different dextrans functionalized with fluorogenic probes, suggesting a glycomonomer oxidation-state dependent mechanism for endosomal uptake and release. Further, transformation of proteins with pendant dextrans endowed increased cellular internalization as assessed with proteins that initiate cell death upon internalization. The structure of glycosylated human ribonuclease 1 afforded insight into the origin of their variable catalytic and thermostability. These investigations unveiled a new helix-capping motif, stemming from N-glycosylation at the C terminus of an α-helix.</Abstract>
<Abstract>Finally, the characterization of cellular reactivity and the associated mass transport properties of SNO-OCT reagents, their ability to engage in triple ligations, and their utility for metabolic labeling experiments were also investigated.</Abstract>
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