title: Leukotriene BLT2 receptor monomers activate the G(i2) GTP-binding protein more efficiently than dimers.
abstract: Accumulating evidence indicates that G protein-coupled receptors can assemble as dimers/oligomers but the role of this phenomenon in G protein coupling and signaling is not yet clear. We have used the purified leukotriene B(4) receptor BLT2 as a model to investigate the capacity of receptor monomers and dimers to activate the adenylyl cyclase inhibitory G(i2) protein. For this, we overexpressed the recombinant receptor as inclusion bodies in the Escherichia coli prokaryotic system, using a human alpha(5) integrin as a fusion partner. This strategy allowed the BLT2 as well as several other G protein-coupled receptors from different families to be produced and purified in large amounts. The BLT2 receptor was then successfully refolded to its native state, as measured by high-affinity LTB(4) binding in the presence of the purified G protein G alpha(i2). The receptor dimer, in which the two protomers displayed a well defined parallel orientation as assessed by fluorescence resonance energy transfer, was then separated from the monomer. Using two methods of receptor-catalyzed guanosine 5'-3-O-(thio)triphosphate binding assay, we clearly demonstrated that monomeric BLT2 stimulates the purified G alpha(i2) beta(1) gamma(2) protein more efficiently than the dimer. These data suggest that assembly of two BLT2 protomers into a dimer results in the reduced ability to signal.
authors.affiliation: CNRS UMR 5203, Institut de Génomique Fonctionnelle, Département de Pharmacologie Moléculaire, Montpellier, France.
authors.forename: Laure
authors.initials: L
authors.lastname: Arcemisbéhère
authors.forename: Tuhinadri
authors.initials: T
authors.lastname: Sen
authors.forename: Laure
authors.initials: L
authors.lastname: Boudier
authors.forename: Marie-Noëlle
authors.initials: MN
authors.lastname: Balestre
authors.forename: Gérald
authors.initials: G
authors.lastname: Gaibelet
authors.forename: Emilie
authors.initials: E
authors.lastname: Detouillon
authors.forename: Hélène
authors.initials: H
authors.lastname: Orcel
authors.forename: Christiane
authors.initials: C
authors.lastname: Mendre
authors.forename: Rita
authors.initials: R
authors.lastname: Rahmeh
authors.forename: Sébastien
authors.initials: S
authors.lastname: Granier
authors.forename: Corinne
authors.initials: C
authors.lastname: Vivès
authors.forename: Franck
authors.initials: F
authors.lastname: Fieschi
authors.forename: Marjorie
authors.initials: M
authors.lastname: Damian
authors.forename: Thierry
authors.initials: T
authors.lastname: Durroux
authors.forename: Jean-Louis
authors.initials: JL
authors.lastname: Banères
authors.forename: Bernard
authors.initials: B
authors.lastname: Mouillac
mesh_terms: D031541:Fluorescence Resonance Energy Transfer; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D044387:GTP-Binding Protein beta Subunits / Q000378:metabolism; D044388:GTP-Binding Protein gamma Subunits / Q000378:metabolism; D006801:Humans; D039564:Integrin alphaV; D011485:Protein Binding; D055503:Protein Multimerization; D018102:Receptors, Leukotriene B4 / Q000502:physiology*; D015398:Signal Transduction
chemical_list: D044387:GTP-Binding Protein beta Subunits; D044388:GTP-Binding Protein gamma Subunits; D039564:Integrin alphaV; C485878:LTB4R2 protein, human; D018102:Receptors, Leukotriene B4; D019206:GTP-Binding Protein alpha Subunits, Gi-Go