Biochemical Assays for Ghrelin Acylation and Inhibition of Ghrelin O-Acyltransferase
- PMID: 31152408
- DOI: 10.1007/978-1-4939-9532-5_18
Biochemical Assays for Ghrelin Acylation and Inhibition of Ghrelin O-Acyltransferase
Abstract
Ghrelin O-acyltransferase (GOAT) is an enzyme responsible for octanoylating and activating ghrelin, a peptide hormone that plays a key role in energy regulation and hunger signaling. Due to its nature as an integral membrane protein, GOAT has yet to be purified in active form which has complicated biochemical and structural studies of GOAT-catalyzed ghrelin acylation. In this chapter, we describe protocols for efficient expression and enrichment of GOAT in insect cell-derived microsomal fraction, HPLC-based assays for GOAT acylation activity employing fluorescently labeled peptides, and assessment of inhibitor potency against GOAT.
Keywords: Enzyme inhibition; Ghrelin; Ghrelin O-acyltransferase (GOAT); HPLC; Membrane-bound O-acyltransferase (MBOAT); Protein acylation; Protein octanoylation.
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