Access Type

Open Access Dissertation

Date of Award

January 2018

Degree Type

Dissertation

Degree Name

Ph.D.

Department

Chemistry

First Advisor

Jennifer L. Stockdill

Abstract

"Conotoxins are mini proteins isolated from marine snails of the Conus family. They have shown activity in the selective inhibition of various ion channels. Therefore, they present an opportunity as possible therapy for a myriad of neurological ailments. The synthesis of conotoxins would enable both biological studies and evaluation of these structures as lead targets for therapeutics. However, establishing a reliable method to consistently folding disulfide rich proteins, including conotoxin, into their native structure has been challenging over few decades. In this research project, it is shown the progress towards a mild, efficient, and broadly applicable methods to generate the desired pattern of disulfide linkages in synthetic polypeptides.

A -allyl chemistry mediated deprotection conditions for Allocam group was optimized to overcome the limitations which prevent its general use in the field. A one-pot approach with Pd-mediated deprotection-oxidation was developed to form disulfide bonds on resin. The utility of the method was evaluated by solid phase synthesis and folding of the carboxy-oxytocin. In addition, 4/7-conotoxin LvIA was synthesized to demonstrate the applicability of novel Pd-DMSO mediated simultaneous Allocam removal and oxidation condition to access more complex disulfide - rich peptides. Two routes were introduced based on position of the Allocam protected cysteines and selection of compatible other cysteine protecting groups for each of the synthetic routes.

The potential of Ellman's reagent as a colorimetric and mild oxidizing agent to form on-resin disulfide bonds in peptides was evaluated. The reaction procedure is simple, no complex time consuming reaction set up is needed and the reaction can be monitored easily by observing the color change of the resin. The colorimetric behavior of Ellman's reagent in different solvents were examined. Ellman's reagent - mediated disulfide bond formation in DMF is relatively slower while it shows faster reaction rates in NMP. This new colorimetric method was utilized in the synthesis of 4/7-conotoxin LvIA and -conotoxin ImI and obtained excellent overall yields.

For the first time, a regioselective on-resin synthesis of three disulfide linkages was achieved by synthesizing linaclotide with careful orthogonal combination of the new Allocam cleavage - oxidation method and Ellman's reagent - mediated on-resin disulfide bond formation method."

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