Synthetic protease-activated class B GPCRs.
Biochem Biophys Res Commun
; 530(1): 246-251, 2020 09 10.
Article
en En
| MEDLINE
| ID: mdl-32828294
ABSTRACT
G-protein coupled receptors (GPCRs) are the ligand detection machinery of a majority of extracellular signaling systems in metazoans. Novel chemical and biological tools to probe the structure-function relationships of GPCRs have impacted both basic and applied GPCR research. To better understand the structure-function of class B GPCRs, we generated receptor-ligand fusion chimeric proteins that can be activated by exogenous enzyme application. As a prototype, fusion proteins of the glucagon-like peptide-1 receptor (GLP-1R) with GLP-1(7-36) and exendin-4(1-39) peptides incorporating enterokinase-cleavable N-termini were generated. These receptors are predicted to generate fusion protein neo-epitopes upon proteolysis with enterokinase that are identical to the N-termini of GLP-1 agonists. This system was validated by measuring enterokinase-dependent GLP-1R mediated cAMP accumulation, and a structure-activity relationship for both linker length and peptide sequence was observed. Moreover, our results show this approach can be used in physiologically relevant cell systems, as GLP-1R-ligand chimeras were shown to induce glucose-dependent insulin secretion in insulinoma cells upon exposure to enterokinase. This approach suggests new strategies for understanding the structure-function of peptide-binding GPCRs.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Péptido Hidrolasas
/
Ingeniería de Proteínas
/
Péptido 1 Similar al Glucagón
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Receptor del Péptido 1 Similar al Glucagón
/
Exenatida
Límite:
Animals
/
Humans
Idioma:
En
Revista:
Biochem Biophys Res Commun
Año:
2020
Tipo del documento:
Article