Taute, Helena2026-01-282026-01-282019-04-052018-07*A2019http://hdl.handle.net/2263/107664Dissertation (MSc)--University of Pretoria, 2018.Antimicrobial peptides (AMPs), small cationic molecules possessing a broad spectrum of bioactivities, have for long been considered promising multi-therapeutic candidates and antibiotic alternatives. High production costs and stability issues have however, limited the application of these molecules, emphasising the need for smaller, more stable, peptide analogues. Such synthetic analogues require structural and functional evaluation prior to their development and use. Previously, the anti-endotoxin and anti-inflammatory potential of synthetic peptides, Os and its cysteine (Cys)-deficient analogue, Os-C, shown to exhibit more potent bactericidal activity than that of their parent peptide, tick defensin OsDef2, had been evaluated. Interestingly, Os demonstrated significantly greater anti-inflammatory activity compared with Os-C. To determine whether the Cys residues or a modified amino acid sequence was responsible for the differences observed between Os and Os-C, current research aimed to determine the cellular, antioxidant, and anti-inflammatory activity of the serine (Ser)-substituted Os analogue, Os(C→S). Selection of Ser was based on structural and chemical similarities between Ser and Cys residues. Neither Os nor Os(C→S) elicited any significant toxic effect against mammalian L929 cells, human erythrocytes or human leukocytes. The cell-penetrating capacity of Os, compared with Os(C→S), although limited appeared to be associated with the presence of Cys residues, shown to be implicated in the overall hydrophobicity of the peptide. In chemico, Os and Os(C→S) were capable of inhibiting free radical production using both electron transfer (ET)- and hydrogen atom transfer (HAT)-based reactions. In vitro, however, the protective effect of Os(C→S) against 2,2′-azobis(2-methylpropionamidine) dihydrochloride (AAPH)-induced oxidative damage was largely negligible, implying some functional contribution of Cys residues to the antioxidant activity of Os. The in chemico nitric oxide (NO) scavenging potential of Os was significantly greater than that of Os(C→S), while ex vivo, extensive platelet activation was observed in whole blood (WB) exposed The absence of significant antioxidant and anti-inflammatory activity by Os(C→S) correlates with the peptide's low hydrophobicity and associated limitation in it’s ability to translocate across the cell membrane, reducing its intracellular availability and subsequent exertion of its protective effects. The thiol groups of sulphur-containing Cys residues present in Os therefore, play a significant role in the cell-penetrating and antioxidant activity of the peptide. Despite this, Os(C→S) may still be considered as an effective AMP owing to its potent antibacterial activity and lack of cytotoxicity in mammalian cells - emphasising the peptide's specificity towards bacterial cells. to both Os and Os(C→S), in the presence and absence of lipopolysaccharide (LPS), consistent with the inability of peptide to bind LPS. The absence of significant antioxidant and anti-inflammatory activity by Os(C→S) correlates with the peptide's low hydrophobicity and associated limitation in it’s ability to translocate across the cell membrane, reducing its intracellular availability and subsequent exertion of its protective effects. The thiol groups of sulphur-containing Cys residues present in Os therefore, play a significant role in the cell-penetrating and antioxidant activity of the peptide. Despite this, Os(C→S) may still be considered as an effective AMP owing to its potent antibacterial activity and lack of cytotoxicity in mammalian cells - emphasising the peptide's specificity towards bacterial cells.en© 2024 University of Pretoria. All rights reserved. The copyright in this work vests in the University of Pretoria. No part of this work may be reproduced or transmitted in any form or by any means, without the prior written permission of the University of Pretoria.UCTDAntimicrobial peptidesDefensinsAntioxidantLipopolysacchariclePlatelet activationDissertation12013685N/A