Broad-spectrum chemokine-binding peptides from class A evasins

Thesis

The chemokine network comprises 46 secreted chemokines that signal through 18 GPCRs, resulting in leukocyte recruitment in inflammation. Despite being a validated therapeutic target, chemokine network redundancy thwarts pharmacological intervention. Class A and Class B tick salivary proteins (evasins) bind CC and CXC-class chemokines, respectively, overcoming redundancy and allowing prolonged blood feeding. Evasins are unsuitable for therapeutic use due to potential immunogenicity and chemokine-class specificity requiring both evasin classes to be used. Peptides previously isolated from the chemokine-binding region of class A evasins inhibit a limited number of CC-class chemokines but lack the breadth of binding needed to overcome redundancy. This thesis tests the hypothesises that phage display can identify and improve peptides from class A evasins with broad chemokine binding and inhibitory activity. By screening a phage-display library from 21 class A evasins, peptides that surprisingly bind both CC and CXC chemokine classes were identified. An exemplar peptide, HD2, from evasin EVA4, inhibited both CC and CXC- class chemokines. Alanine-scanning mutagenesis of HD2 identified critical residues important for inhibitory activity, which correlated with binding activity in phage display. HD845 is a peptide derived from the evasin EV672. HD845 binds and inhibits both CC and CXC-chemokines in phage display and in chemotaxis assays. Saturation mutagenesis of HD845 identified several single mutations that show enhanced chemokine binding and inhibitory activity. Lastly, HD2 served as a template to identify improved mutant peptides capable of inhibiting an atherosclerosis-relevant chemokine pool. These results support the hypothesis that peptides with broad-spectrum chemokine-binding activity can be identified, and their binding affinity and potency improved using phage display.

Authors: Vales, S

• DOI: 10.5287/ora-eo4rq5xry
• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: University of Oxford
• Language: English
• Keywords: inflammation; phage display; evasins; peptide; chemokine
• Subjects: Inflammation; Chemokine