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dc.contributor.authorBecher, Frederik
dc.date.accessioned2021-11-13T03:51:36Z
dc.date.available2021-11-13T03:51:36Z
dc.date.submitted2020-11-02
dc.identifier.urihttps://www.repository.cam.ac.uk/handle/1810/330639
dc.description.abstractBiopharmaceuticals, including proteins and peptides, are becoming increasingly important as therapeutic agents. However, the clinical use of protein and peptide therapeutics is still restricted due to undesirable properties such as their ability to self-assemble and aggregate. These properties not only influence production processes and storage but can also have adverse immunogenic effects within humans. To enhance their use, it is of great importance to understand and be able to manipulate their aggregation behaviour. In vivo, protein- and peptide-based drugs suffer from different problems, such as short lifetimes and low stability. Lipidation of peptides is widely established as a means of increasing stability in vivo. However, relatively little is known about the effect of lipidation on peptide self-assembly and other aggregation phenomena in vitro. The aim of this study is to develop a better understanding of the mechanism of aggregation and amyloid fibrillation of the therapeutic peptide GLP-1 and chemically modified forms of GLP-1. GLP-1, a metabolic hormone, has the ability to decrease blood sugar levels in a glucose-dependent manner by enhancing the secretion of insulin. Previous measurements of GLP-1 aggregation revealed that, at certain pH values, unusual behaviour is observed that has established that the standard nucleation-polymerization mechanism is insufficient to fully describe the reaction under these conditions. To study the aggregation mechanism of GLP-1 and chemically modified forms of the peptide, aggregation kinetics were measured over a wide range of different conditions. In addition, many biophysical techniques, such as AFM, SEM, far-UV CD, FT-IR, 𝜆max, ANS, ex situ ThT and DLS, were employed to probe the structure, size and properties of species in solution during aggregation. The influence of amidation at the C-terminus of GLP-1 on physical stability was assessed and significant differences to GLP-1 were found. The study of this relatively small modification generates a better understanding of the interdependence of net charge, solubility and secondary structure on the aggregation kinetics. The results of these studies also provide further evidence that peptides belonging to the GLP-1 family can form off-pathway oligomeric species that have a significant impact on the aggregation kinetics. Two lipidated analogues of C-terminally amidated GLP-1 (Am-GLP-1) were also studied and the results analysed in detail and compared to those obtained for GLP-1 and Am-GLP-1. Both lipidated peptides show a strong, nearly switch-like, pH dependence. Surprisingly, it was also shown that the amidation of the C- terminus had a bigger influence on the secondary structure of the peptides in comparison to the lipidation.
dc.description.sponsorshipPeterhouse and MedImmune (now AstraZeneca, reference number RG77711)
dc.rightsAll Rights Reserved
dc.rights.urihttps://www.rioxx.net/licenses/all-rights-reserved/
dc.subjectpeptides
dc.subjectlipids
dc.subjectpeptide aggregation
dc.subjectlipidation
dc.subjectGlucagon-like peptide -1
dc.subjectGLP-1
dc.subjectbiopharmaceuticals
dc.subjectThT-assay
dc.titleStudies of the Physical Stability of GLP-1 & Chemically Modified Forms of GLP-1
dc.typeThesis
dc.type.qualificationlevelDoctoral
dc.type.qualificationnameDoctor of Philosophy (PhD)
dc.publisher.institutionUniversity of Cambridge
dc.identifier.doi10.17863/CAM.78083
rioxxterms.licenseref.urihttps://www.rioxx.net/licenses/all-rights-reserved/
rioxxterms.typeThesis
dc.publisher.collegePeterhouse
dc.type.qualificationtitlePhD
cam.supervisorJackson, Sophie
cam.supervisorGomes dos Santos, Ana


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