Date of Award

August 2017

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Chemistry

Advisor(s)

James L. Hougland

Second Advisor

Dacheng Ren

Keywords

Acylation, Enzymology, Ghrelin, Inhibitor, Membrane, Mutagenesis

Subject Categories

Physical Sciences and Mathematics

Abstract

Ghrelin is a circulating peptide hormone that plays a regulatory role in many physiological processes related to energy balance and metabolism. To be recognized by its receptor, growth hormone secretagogue receptor 1a (GHSR1a), the serine 3 residue of ghrelin must be octanoylated. This unique posttranslational modification is catalyzed by the enzyme ghrelin O-acyltransferase (GOAT), a topologically complex integral membrane protein. GOAT is an intriguing target for biochemical studies, as no other enzyme is known to catalyze octanoylation of a serine hydroxyl group. It is also an attractive target for diseases related to ghrelin signaling, such as diabetes and obesity, as blocking GOAT activity would decrease the levels of acylated (active) ghrelin. However, much about GOAT remains undefined: its catalytic residues, active site structure, and mechanism are unknown, and assays studying GOAT activity are limited. The lack of structural and mechanistic information regarding GOAT makes the rational design of small molecule inhibitors difficult, and the inability to easily apply GOAT activity assays to a high throughput format has limited the discovery of small molecule GOAT inhibitors. With these challenges in mind, we have made several improvements to GOAT activity assays which increase product stability and enable real-time monitoring of ghrelin acylation. A library screen of small molecules has revealed a novel class of small molecule inhibitors of human GOAT (hGOAT), which act by covalently modifying a functionally essential cysteine residue within the enzyme. Finally, mutagenesis studies have revealed several residues within hGOAT that are required for activity.

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Open Access

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