Uncovering distinct aspects of amino acid metabolism in Fusobacterium nucleatum
department or school: Science, Irving K. Barber Faculty of (Okanagan)
department or school: Chemistry, Department of (Okanagan)
data provider: DSpace
degree campus: UBCO
creator: Darbyshire, Amanda
issued: 2024-05-16T21:39:24Z
issued: 2024
related degree: Doctor of Philosophy - PhD
degree grantor: University of British Columbia
description: Fusobacterium nucleatum is a gram-negative bacterium that is ubiquitous to the human oral cavity and has been associated to different pathologies. This thesis focuses on three distinct enzymatic pathways that are involved in F. nucleatum amino acid metabolism. Chapter 2 and 3 focus on serine synthase, a pyridoxal 5′-phosphate (PLP)-dependent enzyme that catalyzes the reversible reaction of ʟ-cysteine and H₂O to form ʟ-serine and H₂S. The mechanism for this reaction requires an active site aspartic acid residue (D232) to act as a general acid/base catalyst. Hydrophobic residues surrounding D232 increase the basicity of the carboxylate side chain and restrict the range of nucleophilic substrates that can dock into the second binding site. Mutation of these aromatic residues allows for addition of thiol-based nucleophiles to the α-aminoacrylate intermediate of the PLP cofactor, thus allowing serine synthase to act as a potential biocatalytic tool for noncanonical amino acid synthesis. Chapter 4 focuses on lanthionine synthase, another PLP-dependent enzyme, which catalyzes the condensation reaction of ʟ-cysteine with a second equivalent of ʟ-cysteine to produce ʟ,ʟ-lanthionine and H₂S. In this chapter, we utilized the relaxed substrate scope of lanthionine synthase to produce different lanthionine analogs and test potential inhibitors. We also determined that the physiological production of meso-lanthionine in F. nucleatum is likely through the reaction of lanthionine synthase with ʟ-cysteine and D-cysteine and not through an epimerase. Fn1732 in the F. nucleatum genome may act as a racemase on an amino acid other than ʟ,ʟ- lanthionine. Chapter 5 investigates a putative metallochaperone system for lysine 5,6- aminomutase (5,6-LAM). 5,6-LAM is a PLP- and adenosylcobalamin (AdoCbl)-dependent enzyme involved in the second step of the lysine fermentation pathway, catalyzing the conversion of β-ʟ-lysine to erythro-3,5-diaminohexanoate. Like other AdoCbl-dependent enzymes, 5,…
aggregated CHO: https://circle.library.ubc.ca/rest/handle/2429/88331
note: Uncovering Distinct Aspects of Amino Acid Metabolism in Fusobacterium nucleatum by Amanda Darbyshire B.Sc., The University of British Columbia (Okanagan), 2019 A THESIS SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY in THE COLLEGE OF GRADUATE STUDIES (Biochemistry and Molecular Biology) THE UNIVERSITY OF BRITISH COLUMBIA (Okanagan) May 2024 ã Amanda Darbyshire ii The following individuals certify that they have read, and recommend to the College of Graduate Studies for acceptance, a thesis/dissertation entitled: Uncovering Distinct Aspects of Amino Acid Metabolism in Fusobacterium nucleatum submitted by Amanda Darbyshire in partial fulfillment of the requirements of the degree of Doctor of Philosophy in Biochemistry and Molecular Biology Dr. Kirsten Wolthers (Irving K. Barber Faculty of Science) Supervisor Dr. Frederic Menard (Irving K. Barber Faculty of Science) Supervisory Committee Member Dr. Thu-Thuy Dang (Irving K. Barber Faculty of Science) Supervisory Committee Member Dr. Natalie Strynadka (Faculty of Medicine) University Examiner Dr. Stephen Bearne (Dalhousie University) External Examiner iii Abstract Fusobacterium nucleatum is a gram-negative bacterium that is ubiquitous to the human oral cavity and has been associated to different pathologies. This thesis focuses on three distinct enzymatic pathways that are involved in F. nucleatum amino acid metabolism. Chapter 2 and 3 focus on serine synthase, a pyridoxal 5′-phosphate (PLP)-dependent enzyme that catalyzes the reversible reaction of L-cysteine and H2O to form L-serine and H2S. The mechanism for this reaction requires an active site aspartic acid residue (D232) to act as a general acid/base catalyst. Hydrophobic residues surrounding D232 increase the basicity of the carboxylate side chain and restrict the range of nucleophilic substrates that can dock into the second binding site. Mutation of these aromatic residues allows for addition of thiol-based nucleophil…
has type: Thesis/Dissertation
date issued: 2024-09
is shown at: 10.14288/1.0443562
language: eng
degree discipline: Biochemistry and Molecular Biology
provider: Vancouver : University of British Columbia Library
publisher: University of British Columbia
rights: Attribution-NonCommercial-NoDerivatives 4.0 International
rights URI: http://creativecommons.org/licenses/by-nc-nd/4.0/
scholar level: Graduate
contributor: Wolthers, Kirsten
type: Text
identifier URI: http://hdl.handle.net/2429/88331