Date of Award

2020

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Drug Discovery and Biomedical Sciences

College

College of Graduate Studies

First Advisor

Frank Mari

Second Advisor

Mark Hamann

Third Advisor

Patrick Woster

Fourth Advisor

Christopher Davies

Fifth Advisor

Yuri Peterson

Abstract

Cone snail venom is a mixture of disulfide-constrained peptides (conotoxins), hormone-like peptides, and proteins that have been ‘weaponized’ for predation and defense. Venom peptides, or conopeptides, have efficiently evolved to bind receptors and ion channels that modulate the neuromuscular, cardiovascular, and central nervous systems in prey species. With over 850 species of cone snails, each with unique venom concoctions, cone snail venom is a valuable source of novel pharmacological probes and potential drug leads. However, the complexity of the venom poses a challenge for drug discovery. Contributing to the complexity is 1) a wide range in molecular weight 2) peptide hyper-variability by post-translational modifications and 3) many potential molecular targets to pursue. In this research, a ‘venomics’ approach was employed for the global identification of venom components. This ‘venomics’ methodology combines RNAseq data from the venom duct and proteomic data from raw injected venom to identify novel conopeptides. This project was a data-driven effort to define the venom components of the cone snail, Conus purpurascens, and to stimulate further hypothesis-driven studies. First, 21 new base conopeptides were identified from the injected venom of Conus purpurascens, a fish-hunting cone snail native to the Pacific coast of Central America. The molecular targets were projected based on homology to previously characterized conopeptides. The newly identified conopeptides included α-conotoxin, α-PID. Alpha-conotoxins are inhibitory ligands of nicotinic acetylcholine receptors (nAChRs), and the most ubiquitous venom components across the Conus genus. Ligands of nAChRs are clinically important for addiction, cognitive disorders, neurodegenerative diseases, and pain. Functional characterization of α-PID and three other α-conotoxins was performed to test their activity on different nAChR subtypes using heterologous receptor expression and molecular modeling techniques. A unique insulin-like peptide (Con-Ins P1) was also identified and was the first instance of an insulin-like peptide identified directly from injected venom. This research demonstrates how discovery-based ‘venomics’ workflows can be used to yield novel peptides with pharmacological applications and stimulate further hypothesis-driven experiments.

Rights

All rights reserved. Copyright is held by the author.

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