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Peptide Toxin Bioengineering: Exploring the Potential of Cyclized Conotoxins as Oral Drugs, Fluorescent Probes and Phyla - Selective Peptides
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|Title:||Peptide Toxin Bioengineering: Exploring the Potential of Cyclized Conotoxins as Oral Drugs, Fluorescent Probes and Phyla - Selective Peptides|
Native Chemical Ligation
show 1 moreHuwentoxin and Bioengineering
|Date Issued:||Dec 2015|
|Publisher:||[Honolulu] : [University of Hawaii at Manoa], [December 2015]|
|Abstract:||Studies into N- to C-terminal cyclic peptide backbone structures have provided for the lateral transition of important principles and strategies that clearly resonate within the world of bioactive peptides and peptide toxins. The ability to transform peptide biologics into stable and orally active constituents represents a major pharmacological achievement. This progression has been forthcoming and is potentially intensified by the diminishing expectations of current small organic molecule pipelines. |
For our studies we have decided to use conotoxins as the model natural product for cyclization. Conotoxins are disulfide rich neurotoxins found in the venoms of marine cone snails. These neuropeptides have a high number of post-translational modifications (PTMs), which manifest itself in phyla specificity and receptor selectivity. Conotoxins are typically about 10-40 amino acids in length, but exhibit many secondary motifs such as α-helices and β-sheets that are normally found in larger proteins. Their high level of selectivity, potency and structural integrity make them prime candidates for oral drug models, drug scaffolds and bio-conjugation tools. Venom derived peptides from marine cone snails, conotoxins, have demonstrated unique pharmacological targeting properties that have been pivotal in advancing medical research. The awareness of their true toxic origins and potent pharmacological nature is emphasized by their ‘select agent’ classification by the US Centers for Disease Control and Prevention.
In order to cyclize these natural peptides, we will adapt the Native Chemical Ligation (NCL) strategy. Current techniques use Hydrogen fluoride (HF) to create the thioester, which can ultimately be used for native chemical ligation/cyclization. The use of HF is not recommended due to its highly hazardous and toxic nature. Furthermore, since HF requires special apparatus and hood space its application is limited only to a few privileged laboratories. This research aimed at replacing HF with Trifluoromethanesulfonic acid (TFMSA) without compromising with the yield of the reaction. In order to optimize the reaction conditions to achieve target concentration various variables such as time, acid concentration, and different orthogonal protection group scavengers will be examined. By replacing HF with TFMSA, which is less toxic and uses conventional lab ware, we aim to make the technique of thioester ligation/cyclization safer and accessible. Here we use a 33 amino acid (αα) test peptide, Huwentoxin-I (HwTx-I) as a candidate to test this novel protocol. Structurally HwTx-I has 6 individual Cysteines (Cys) and an X—Cys-Cys—X sequence mid-region, which makes it an ideal candidate for thioester ligation.
Once we have established a protocol for cyclization/ligation, this novel protocol will be used to synthesize the cyclic conopeptide. After synthesis we will conduct stability and bioactivity assay on the cyclic peptide to assess the impact of cyclization on bioactivity and stability. Cyclized conotoxins display increased structural stability and resistance to enzymatic degradation and may be used to create orally active conotoxin therapeutics. In all, the works presented in this dissertation display a progression in conotoxin bioengineering to improve their pharmacological properties so they may be developed as therapeutics, biopesticides or bioconjugation tools.
|Description:||Ph.D. University of Hawaii at Manoa 2015.|
Includes bibliographical references.
|Appears in Collections:||
Ph.D. - Molecular Biosciences and Bioengineering|
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