Chronic pain affects almost 10 million people in the UK but despite this, few effective treatments exist. Research has shown that targeting the Nav1.7 ion channel provides a novel approach to treatment. ProTx-II, a 30 amino acid peptide isolated from Tarantula venom, is highly selective for the channel (IC50 value - 0.3nm in vitro) but in vivo results were less promising. ProTx-II contains three interlocking disulfide bonds connected in a distinctive pattern. To investigate the structure-activity relationship between the peptide and the ion channel, truncated analogues based on the individual cysteine rings were synthesised. This thesis investigates the effect of replacing the disulfide bond with a thioether linkage through the incorporation of a novel diastereoisomer of the non-natural amino acid lanthionine to produce hydrolytically stable compounds. Lanthionine can be thought of as two alanine residues connected by a thioether linkage at the β-carbon. Improvements in the synthesis allowed for the first large scale production of lanthionine enabling multiple attempts at peptide synthesis to be carried out. Work was also begun on the synthesis of cystathionine, a second thioether linker containing an extra CH2 group between the amino acid centres. Novel methodologies were investigated for the incorporation of a new diastereomer of lanthionine containing two L-amino acid centres using a new microwave-based methodology. This methodology has so far produced the largest known synthetic rings containing a lanthionine bridge, along with the first interlocking disulfide and thioether bridged compounds. The analogues produced were modelled using in silico mutagenesis techniques and the effect of these analogues on the hNav1.7 receptor was investigated using an automated patch clamp assay. Finally, analysis of the connectivity of ProTx-II was carried out using a combination of digestion and MS/MS experiments.
|Creators||Wright, Z. V. F.|
|Publisher||University College London (University of London)|
|Source Sets||Ethos UK|
|Type||Electronic Thesis or Dissertation|
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