2013
High-Level Expression, Purification and Characterization of a Constitutively Active Thromboxane A2 Receptor Polymorphic Variant
Xu B, Chakraborty R, Eilers M, Dakshinamurti S, O’Neil J, Smith S, Bhullar R, Chelikani P. High-Level Expression, Purification and Characterization of a Constitutively Active Thromboxane A2 Receptor Polymorphic Variant. PLOS ONE 2013, 8: e76481. PMID: 24086743, PMCID: PMC3781061, DOI: 10.1371/journal.pone.0076481.Peer-Reviewed Original ResearchConceptsG protein-coupled receptorsHigh-level expressionThromboxane A2 receptorSuccessful high-level expressionCell linesHigh-resolution crystal structuresProper expression systemMammalian cell linesResolution crystal structureProtein-coupled receptorsSingle-step affinity purificationStep affinity purificationCircular dichroism spectropolarimetryMembrane proteinsActive mutantAffinity purificationExpression systemBiophysical characterizationConstitutive signalingT geneSecondary structure changesT mutantGenetic variantsDiffusible ligandsHomogeneous glycosylationNew Insights into Structural Determinants for Prostanoid Thromboxane A2 Receptor- and Prostacyclin Receptor-G Protein Coupling
Chakraborty R, Pydi SP, Gleim S, Bhullar RP, Hwa J, Dakshinamurti S, Chelikani P. New Insights into Structural Determinants for Prostanoid Thromboxane A2 Receptor- and Prostacyclin Receptor-G Protein Coupling. Molecular And Cellular Biology 2013, 33: 184-193. PMID: 23109431, PMCID: PMC3554117, DOI: 10.1128/mcb.00725-12.Peer-Reviewed Original ResearchMeSH KeywordsAmino Acid SequenceBinding SitesCalciumFluorescent Antibody TechniqueGTP-Binding ProteinsHEK293 CellsHumansInositol 1,4,5-TrisphosphateLigandsModels, MolecularMolecular Sequence DataMutagenesis, Site-DirectedNonlinear DynamicsProtein BindingProtein ConformationReceptors, EpoprostenolReceptors, Thromboxane A2, Prostaglandin H2Signal TransductionConceptsG protein-coupled receptorsG protein specificityThromboxane A2 receptorProtein interactionsProtein specificityIntracellular loopChimeric receptorsG proteinsGPCR-G protein interactionsProtein couplingHeterotrimeric G proteinsCognate G-proteinSite-directed mutagenesisNew molecular insightsProtein-coupled receptorsG protein couplingReceptor-G protein interactionG protein activationA2 receptorsReceptor-G protein couplingStructural basisMolecular insightsProtein activationMolecular natureStructural determinants
2012
Role of rhodopsin N-terminus in structure and function of rhodopsin-bitter taste receptor chimeras
Pydi S, Chakraborty R, Bhullar R, Chelikani P. Role of rhodopsin N-terminus in structure and function of rhodopsin-bitter taste receptor chimeras. Biochemical And Biophysical Research Communications 2012, 430: 179-182. PMID: 23159609, DOI: 10.1016/j.bbrc.2012.11.029.Peer-Reviewed Original ResearchConceptsG protein-coupled receptorsBitter taste receptorsTaste receptorsProtein-coupled receptorsBitter taste sensationReceptor expressionLow-level expressionCalcium imagingReceptor functionFlow cytometryReceptorsTaste sensationAllosteric ligand bindingT2RsReceptor chimerasAmino acidsExpressionReceptor structureN-terminal 33N-terminal 33 amino acidsN-terminal sequenceCytometrySite-Directed Mutations and the Polymorphic Variant Ala160Thr in the Human Thromboxane Receptor Uncover a Structural Role for Transmembrane Helix 4
Chakraborty R, Pydi SP, Gleim S, Dakshinamurti S, Hwa J, Chelikani P. Site-Directed Mutations and the Polymorphic Variant Ala160Thr in the Human Thromboxane Receptor Uncover a Structural Role for Transmembrane Helix 4. PLOS ONE 2012, 7: e29996. PMID: 22272267, PMCID: PMC3260207, DOI: 10.1371/journal.pone.0029996.Peer-Reviewed Original ResearchMeSH Keywords15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic AcidAlanineAmino Acid SequenceAnimalsBinding SitesBinding, CompetitiveBridged Bicyclo Compounds, HeterocyclicCalciumChlorocebus aethiopsCOS CellsFatty Acids, UnsaturatedGlycineHEK293 CellsHumansHydrazinesMicroscopy, FluorescenceModels, MolecularMolecular Sequence DataMutagenesis, Site-DirectedMutant ProteinsMutation, MissensePolymorphism, Single NucleotideProtein Structure, SecondaryRadioligand AssayReceptors, Thromboxane A2, Prostaglandin H2Structure-Activity RelationshipTemperature