Discovery of new glycoside phosphorylases through rational, genomic and metagenomic exploration
department or school: Science, Faculty of
data provider: DSpace
degree campus: UBCV
creator: Macdonald, Spencer
issued: 2022-06-30T07:00:00Z
issued: 2021
related degree: Doctor of Philosophy - PhD
degree grantor: University of British Columbia
description: Carbohydrates are abundant throughout nature, play key roles in diverse biological processes and form fundamental components of a wide range of functional materials. Their inherent complexity is a double edge sword in that it allows carbohydrates to adopt a diverse range of structures and functions, but in an industrial setting, that complexity can also make them especially difficult and costly to synthesize in a strictly defined manner. To help ensure uniform production, carbohydrate manufacturing processes are known to utilize Carbohydrate Active Enzymes (CAZymes), which have innate substrate specificities and conformational control. A class of CAZymes, known as glycoside phosphorylases (GPs) catalyze the reversible phosphorolysis of glycosidic bonds, releasing sugar 1-phosphates and have considerable potential as catalysts for the assembly of useful carbohydrates for products ranging from functional foods and prebiotics to novel materials. However, the substrate diversity of currently identified GPs is relatively small, limiting their practical applications. To address this limitation, I used a combined approach drawing upon rational, metagenomic and genomic exploration methods to discover new GPs. First, using a rational approach, I identified a new GP activity, the first reported with a β-retaining mechanism, while investigating a mechanistic oddity observed in the GH3 CAZy family. Next, I developed and deployed two metagenomic screening methodologies that targeted GP activity in the vast reservoir of uncultivated genetic diversity encoded in microbial communities inhabiting natural and engineered ecosystems. This approach yielded eight new GPs and established a screening paradigm that can be applied to an even greater range of GP activities. Lastly, a phylogenomically diverse, synthetic gene GP library was characterized, which led to the discovery of another previously undiscovered GP activity, a new biopolymer (that I named acholetin), and established a high …
aggregated CHO: https://circle.library.ubc.ca/rest/handle/2429/78887
note: Discovery of New GlycosidePhosphorylases through Rational,Genomic and Metagenomic ExplorationbySpencer MacdonaldB.Sc., Wilfrid Laurier University, 2010M.Sc., The University of British Columbia, 2015A THESIS SUBMITTED IN PARTIAL FULFILLMENT OFTHE REQUIREMENTS FOR THE DEGREE OFDOCTOR OF PHILOSOPHYinThe Faculty of Graduate and Postdoctoral Studies(Genome Science and Technology)THE UNIVERSITY OF BRITISH COLUMBIA(Vancouver)July 2021© Spencer Macdonald 2021The following individuals certify that they have read, and recommend to the Faculty of Graduateand Postdoctoral Studies for acceptance, the dissertation entitled:Discovery of New Glycoside Phosphorylases throughRational, Genomic and Metagenomic ExplorationSubmitted By Spencer Macdonald in partial fulfillment of the requirements forthe degree of Doctor of Philosophyin Genome Science and TechnologyExamining Committee:Stephen G. Withers, Professor, Chemistry and Biochemistry, UBCSupervisorHarry Brummer, Professor, Michael Smith Laboratories, UBCSupervisory Committee MemberKatherine S. Ryan, Associate Professor, Chemistry, UBCUniversity ExaminerOrlando Rojas, Professor, Chemical and Biological Engineering, UBCUniversity ExaminerRobert A. Field, Professor, Chemistry, The University of ManchesterExternal ExaminerAdditional Supervisory Committee Members:Steven J. Hallam, Professor, Microbiology and Immunology, UBCSupervisory Committee MemberLindsay D. Eltis, Professor, Microbiology and Immunology, UBCSupervisory Committee MemberiiAbstractCarbohydrates are abundant throughout nature, play key roles in diverse biological processes andform fundamental components of a wide range of functional materials. Their inherent complexity isa double edge sword in that it allows carbohydrates to adopt a diverse range of structures and func-tions, but in an industrial setting, that complexity can also make them especially difficult and costlyto synthesize in a strictly defined manner. To help ensure uniform production, carbohydrate man-ufactu…
has type: Thesis/Dissertation
date issued: 2021-11
is shown at: 10.14288/1.0400085
language: eng
degree discipline: Genome Science and Technology
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: Withers, Stephen G.
type: Text
identifier URI: http://hdl.handle.net/2429/78887