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dc.contributor.authorLukose, Vinitaen_US
dc.contributor.otherMassachusetts Institute of Technology. Department of Chemistry.en_US
dc.date.accessioned2016-03-03T21:08:19Z
dc.date.available2016-03-03T21:08:19Z
dc.date.copyright2015en_US
dc.date.issued2015en_US
dc.identifier.urihttp://hdl.handle.net/1721.1/101550
dc.descriptionThesis: Ph. D., Massachusetts Institute of Technology, Department of Chemistry, 2015.en_US
dc.descriptionCataloged from PDF version of thesis.en_US
dc.descriptionIncludes bibliographical references.en_US
dc.description.abstractBacterial cell surfaces prominently feature a variety of complex glycoconjugates. Although these structures are very diverse, the biosynthesis of these glycoconjugates shares common themes. In particular, a priming glycosyltransferase or phosphoglycosyltransferase (PGT) initiates the biosynthesis of glycans by transferring a Cl'-phosphosugar onto a polyprenol phosphate substrate. The membrane-bound polyprenol diphosphosugar product is then further elaborated by additional glycosyltransferases, flipped across the inner bacterial membrane, and ultimately used as glycosyl donor substrate in transfer to a final acceptor substrate, which, is a lipid, a protein or another glycan. This thesis focuses on the priming GTs that catalyze the first step in glycoconjugate biosynthesis. This class of enzymes is structurally diverse and includes several distinct families of enzymes with different structures and membrane topologies. PglC, the smallest and structurally, the simplest PGT found in Campylobacter jejuni, represents a tractable model with which to investigate this class of enzymes. First, efforts to structurally characterize PglC are discussed, with a focus on the challenges associated with expressing, purifying and characterizing integral membrane proteins. The next section of this thesis presents the functional characterization of PglC using a variety of biochemical techniques. In addition, bioinformatics analysis of PglC and related PGT families was employed to provide insight into critical residues required for catalytic activity. To complement this, a predicted structure of PgIC was developed, based on the wealth of information contained in the sequences of homologs of PglC. Kinetic analysis of PglC was used to investigate the mechanism of the PGT reaction. Together, these studies aided in the design and evaluation of inhibitors of PgIC, inspired by the nucleoside antibiotics tunicamycin and mureidomycin. In the final chapter, the enzymes in the N-linked protein glycosylation pathway in Campylobacter jejuni were evaluated for their tolerance for azide-modified UDP-sugar substrates. In vitro experiments were employed to investigate the potential of these enzymes for executing the chemoenzymatic synthesis of the C. jejuni glycan with azide-modified sugars at discrete targeted positions. Attempts to metabolically label C. jejuni with azide-modified sugars for in vivo incorporation into the N-glycan are also presented.en_US
dc.description.statementofresponsibilityby Vinita Lukose.en_US
dc.format.extent176 pagesen_US
dc.language.isoengen_US
dc.publisherMassachusetts Institute of Technologyen_US
dc.rightsM.I.T. theses are protected by copyright. They may be viewed from this source for any purpose, but reproduction or distribution in any format is prohibited without written permission. See provided URL for inquiries about permission.en_US
dc.rights.urihttp://dspace.mit.edu/handle/1721.1/7582en_US
dc.subjectChemistry.en_US
dc.titlePriming and processing glycosyltransferases in bacterial N-linked glycosylation pathwaysen_US
dc.typeThesisen_US
dc.description.degreePh. D.en_US
dc.contributor.departmentMassachusetts Institute of Technology. Department of Chemistry.en_US
dc.identifier.oclc940565461en_US


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