Alanine-261 in intracellular loop III of the human gonadotropin-releasing hormone receptor is crucial for G-protein coupling and receptor internalization

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dc.contributor.authorMyburgh D.B.
dc.contributor.authorMillar R.P.
dc.contributor.authorHapgood J.P.
dc.date.accessioned2011-05-15T16:03:23Z
dc.date.available2011-05-15T16:03:23Z
dc.date.issued1998
dc.description.abstractGonadotropin-releasing hormone (GnRH) is a decapeptide that regulates reproductive function via binding to the GnRH receptor, which is a G-protein-coupled receptor (GPCR). For several members of this family, the C-terminal domain of intracellular loop III is important in ligand-mediated coupling to G-proteins; mutations in that region can lead to constitutive activity. A specific alanine residue is involved in certain GPCRs, the equivalent of which is Ala-261 in the GnRH receptor. Mutation of this residue to Leu, Ile, Lys, Glu or Phe in the human GnRH receptor did not result in constitutive activity and instead led to complete uncoupling of the receptor (failure to support GnRH-stimulated inositol phosphate production). When this residue was mutated to Gly, Pro, Ser or Val, inositol phosphate production was still supported. All the mutants retained the ability to bind ligand, and the affinity for ligand, where measured, was unchanged. These results show that Ala261 cannot be involved in ligand binding but is critical for coupling of the receptor to its cognate G-protein. Coupling is also dependent on the size of the residue in position 261. When the amino acid side chain has a molecular mass of less than 40 Da efficient coupling is still possible, but when its molecular mass exceeds 50 Da the receptor is uncoupled. Internalization studies on the Ala261→Lys mutant showed a marked decrease in receptor internalization compared with the wild type, indicating that coupling is necessary for effective receptor internalization in the GnRH receptor system. Activation of protein kinase C (with PMA), but not protein kinase A (with forskolin) markedly increased the internalization of the mutant receptor while having a small effect on the wild-type receptor.
dc.description.versionArticle
dc.identifier.citationBiochemical Journal
dc.identifier.citation331
dc.identifier.citation3
dc.identifier.issn2646021
dc.identifier.urihttp://hdl.handle.net/10019.1/12601
dc.subjectalanine
dc.subjectgonadorelin receptor
dc.subjectgonadotropin
dc.subjectguanine nucleotide binding protein
dc.subjectlysine
dc.subjectprotein kinase c
dc.subjectarticle
dc.subjectcarboxy terminal sequence
dc.subjectgene mutation
dc.subjectinternalization
dc.subjectligand binding
dc.subjectpriority journal
dc.subjectprotein interaction
dc.subjectreproduction
dc.subjectAlanine
dc.subjectAnimals
dc.subjectBinding Sites
dc.subjectBinding, Competitive
dc.subjectCOS Cells
dc.subjectEndocytosis
dc.subjectForskolin
dc.subjectGonadotropin-Releasing Hormone
dc.subjectGTP-Binding Proteins
dc.subjectHumans
dc.subjectInositol Phosphates
dc.subjectLigands
dc.subjectMutagenesis, Site-Directed
dc.subjectProtein Binding
dc.subjectReceptors, LHRH
dc.subjectTetradecanoylphorbol Acetate
dc.subjectTransfection
dc.titleAlanine-261 in intracellular loop III of the human gonadotropin-releasing hormone receptor is crucial for G-protein coupling and receptor internalization
dc.typeArticle
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