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dc.contributor.advisorNaismith, Jim
dc.contributor.authorBent, Andrew F.
dc.coverage.spatialxxv, 236 p.en_US
dc.date.accessioned2017-03-06T11:26:53Z
dc.date.available2017-03-06T11:26:53Z
dc.date.issued2016-06-22
dc.identifieruk.bl.ethos.707258
dc.identifier.urihttp://hdl.handle.net/10023/10404
dc.description.abstractCyclic peptides have potential as scaffolds for novel pharmaceuticals, however their chemical synthesis can be challenging and as such natural sources are often explored. Several species of cyanobacteria produce a family of cyclic peptides, the cyanobactins, through the ribosomal synthesis of precursor peptides and post-translational tailoring. The patellamides, a member of the cyanobactin family, are cyclic octapeptides containing D-stereo centres and heterocyclised amino acids. A single gene cluster, patA - patG, contains the genes for the expression of the precursor peptide and the enzymes responsible for post-translational modifications including a heterocyclase, protease, macrocyclase and oxidase. Biochemical and structural analysis on the patellamide and related cyanobactin pathways has been carried out. The crystal structure of PatF, a proposed prenyl transferase, has been determined, highlighting that it is likely evolutionary inactive due to changes to key residues when compared to active homologues. This is in agreement with the knowledge that no naturally prenylated patellamides have been discovered to date. The crystal structure of the macrocyclase domain of PatG has been determined in complex with a substrate analogue peptide. The structure, together with biochemical analysis has allowed a mechanism of macrocyclisation to be proposed, confirming the requirement of a specific substrate conformation to enable macrocyclisation. Using isolated enzymes from the patellamide and related pathways, a small scale library of macrocycles made up of diverse sequences has been created in vitro and characterised by mass spectrometry and in certain cases NMR. In order to further enhance diversity, macrocycles containing unnatural amino acids have been created using three approaches; SeCys derived precursor peptides, intein-mediated peptide ligation and pEVOL amber codon technology. Finally, two oxidase enzymes from cyanobactin pathways have been purified, characterised and confirmed active for thiazoline oxidation. Native X-ray datasets on crystals of the oxidase CyaGox have been collected and phasing trials are on-going.en_US
dc.language.isoenen_US
dc.publisherUniversity of St Andrews
dc.rightsCreative Commons Attribution-NonCommercial-NoDerivatives 4.0 International
dc.rights.urihttp://creativecommons.org/licenses/by-nc-nd/4.0/
dc.subjectCyanobactinsen_US
dc.subjectCyclic peptidesen_US
dc.subjectX-ray crystallographyen_US
dc.subject.lccQP552.P4B4
dc.subject.lcshCyclic peptidesen
dc.subject.lcshCyclic peptides--Synthesisen
dc.subject.lcshCyanobacteriaen
dc.subject.lcshX-ray crystallographyen
dc.titleStructural and biochemical studies on the biosynthetic pathways of cyanobactinsen_US
dc.typeThesisen_US
dc.contributor.sponsorBiotechnology and Biological Sciences Research Council (BBSRC)en_US
dc.type.qualificationlevelDoctoralen_US
dc.type.qualificationnamePhD Doctor of Philosophyen_US
dc.publisher.institutionThe University of St Andrewsen_US


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