| Summary |
Fibrinogen is a 340 kDa glycoprotein that is capable of creating an insoluble clot during a process called fibrin polymerization. This is initiated with the conversion of fibrinogen to fibrin monomer conversion through cleavage and release of fibrinopeptides A and B, by an enzyme called thrombin. As proof-of-principle, functional studies on the WT protein from commercial fibrinogen were conducted in which the N-linked glycans were removed by PNGase F or processed with neuraminidase. Turbidity and fibrinopeptide release assays in conjunction were employed to characterize the effects of N-linked glycans on fibrin polymerization. In order to fully depict the fibrinogen structure and function of its glycans, a functional expression and purification system is necessary. A recombinant system using transient transfection methods in HEK and CHO cells is being developed with varied conditions such as different PEI:DNA ratios, use of circular versus linear DNA, and performing the transfection with suspended vs adherent cells. A novel synthetic peptide Fmoc-GPRPFPAWK, bound to NHS-activated Sepharose 4 Fast Flow resin is introduced as a viable affinity based purification technique. This will enable future studies linked to site-specific glycans. |
| General note | Presented to the faculty of the Department of Chemistry. |
| General note | Advisor: Adam R. Offenbacher |
| General note | Title from PDF t.p. (viewed February 9, 2022). |
| Dissertation note | M.S. East Carolina University 2021 |
| Bibliography note | Includes bibliographical references. |
| Technical details | System requirements: Adobe Reader. |
| Technical details | Mode of access: World Wide Web. |
| Genre/form | Academic theses. |
| Genre/form | Academic theses. |
