Chemogenomic Analysis of G-Protein Coupled Receptors and Their Ligands Deciphers Locks and Keys Governing Diverse Aspects of Signalling

Understanding the molecular mechanism of signalling in the important super-family of G-protein-coupled receptors (GPCRs) is causally related to questions of how and where these receptors can be activated or inhibited. In this context, it is of great interest to unravel the common molecular features of GPCRs as well as those related to an active or inactive state or to subtype specific G-protein coupling. In our underlying chemogenomics study, we analyse for the first time the statistical link between the properties of G-protein-coupled receptors and GPCR ligands. The technique of mutual information (MI) is able to reveal statistical inter-dependence between variations in amino acid residues on the one hand and variations in ligand molecular descriptors on the other. Although this MI analysis uses novel information that differs from the results of known site-directed mutagenesis studies or published GPCR crystal structures, the method is capable of identifying the well-known common ligand binding region of GPCRs between the upper part of the seven transmembrane helices and the second extracellular loop. The analysis shows amino acid positions that are sensitive to either stimulating (agonistic) or inhibitory (antagonistic) ligand effects or both. It appears that amino acid positions for antagonistic and agonistic effects are both concentrated around the extracellular region, but selective agonistic effects are cumulated between transmembrane helices (TMHs) 2, 3, and ECL2, while selective residues for antagonistic effects are located at the top of helices 5 and 6. Above all, the MI analysis provides detailed indications about amino acids located in the transmembrane region of these receptors that determine G-protein signalling pathway preferences.

[1]  Mark R. Chance,et al.  Structural waters define a functional channel mediating activation of the GPCR, rhodopsin , 2009, Proceedings of the National Academy of Sciences.

[2]  B. Kieffer,et al.  The second extracellular loop: a damper for G protein–coupled receptors? , 2005, Nature Structural &Molecular Biology.

[3]  J. Simms,et al.  Systematic analysis of the entire second extracellular loop of the V(1a) vasopressin receptor: key residues, conserved throughout a G-protein-coupled receptor family, identified. , 2007, The Journal of biological chemistry.

[4]  J. Raymond.  Multiple mechanisms of receptor-G protein signaling specificity. , 1995, The American journal of physiology.

[5]  David E. Gloriam,et al.  Comprehensive repertoire and phylogenetic analysis of the G protein-coupled receptors in human and mouse. , 2006, Genomics.

[6]  Edgar Jacoby,et al.  The 7 TM G‐Protein‐Coupled Receptor Target Family , 2006, ChemMedChem.

[7]  Shay Bar-Haim,et al.  G protein-coupled receptors: in silico drug discovery in 3D. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[8]  Michael T. M. Emmerich,et al.  A novel chemogenomics analysis of G protein-coupled receptors (GPCRs) and their ligands: a potential strategy for receptor de-orphanization , 2010, BMC Bioinformatics.

[9]  T. Schwartz,et al.  Molecular mechanism of 7TM receptor activation--a global toggle switch model. , 2006, Annual review of pharmacology and toxicology.

[10]  T. Gudermann,et al.  The luteinizing hormone receptor activates phospholipase C via preferential coupling to Gi2. , 1999, Biochemistry.

[11]  Konstantin V. Balakin,et al.  Focused chemistry from annotated libraries , 2006 .

[12]  W. Thomas,et al.  Role of helix 8 in G protein-coupled receptors based on structure–function studies on the type 1 angiotensin receptor , 2009, Molecular and Cellular Endocrinology.

[13]  P. Evans,et al.  Agonist‐specific coupling of a cloned Drosophila octopamine/tyramine receptor to multiple second messenger systems. , 1994, The EMBO journal.

[14]  B. Roth,et al.  Potential modes of interaction of 9-aminomethyl-9,10-dihydroanthracene (AMDA) derivatives with the 5-HT2A receptor: a ligand structure-affinity relationship, receptor mutagenesis and receptor modeling investigation. , 2008, Journal of medicinal chemistry.

[15]  Wentian Li.  Mutual information functions versus correlation functions , 1990 .

[16]  G Vriend,et al.  Receptors coupling to G proteins: Is there a signal behind the sequence? , 2000, Proteins.

[17]  Jean-Philippe Vert,et al.  Protein-ligand interaction prediction: an improved chemogenomics approach , 2008, Bioinform..

[18]  L. Pardo,et al.  Ligand-specific regulation of the extracellular surface of a G protein coupled receptor , 2009, Nature.

[19]  Michael G. Lerner,et al.  Binding MOAD (Mother Of All Databases) , 2005, Proteins.

[20]  R. Paschke,et al.  Contacts between Extracellular Loop Two and Transmembrane Helix Six Determine Basal Activity of the Thyroid-stimulating Hormone Receptor* , 2007, Journal of Biological Chemistry.

[21]  Patrick Scheerer,et al.  Crystal structure of the ligand-free G-protein-coupled receptor opsin , 2008, Nature.

[22]  H. Schiöth,et al.  The G-protein-coupled receptors in the human genome form five main families. Phylogenetic analysis, paralogon groups, and fingerprints. , 2003, Molecular pharmacology.

[23]  S. Peluso,et al.  Mapping the binding site of melanocortin 4 receptor agonists: a hydrophobic pocket formed by I3.28(125), I3.32(129), and I7.42(291) is critical for receptor activation. , 2006, Journal of medicinal chemistry.

[24]  T. Sakmar,et al.  Tracking G-protein-coupled receptor activation using genetically encoded infrared probes , 2010, Nature.

[25]  M. Tota,et al.  Molecular determinants of ligand binding to the human melanocortin-4 receptor. , 2000, Biochemistry.

[26]  J. Tyndall,et al.  GPCR agonists and antagonists in the clinic. , 2005, Medicinal chemistry (Shariqah (United Arab Emirates)).

[27]  J. Gutkind,et al.  G-protein-coupled receptors and cancer , 2007, Nature Reviews Cancer.

[28]  Satoshi Niijima,et al.  GLIDA: GPCR—ligand database for chemical genomics drug discovery—database and tools update , 2007, Nucleic Acids Res..

[29]  Steven O. Smith,et al.  Multiple switches in G protein-coupled receptor activation. , 2009, Trends in pharmacological sciences.

[30]  H. Hamm,et al.  Structural and dynamical changes in an α-subunit of a heterotrimeric G protein along the activation pathway , 2006, Proceedings of the National Academy of Sciences.

[31]  B. Kobilka,et al.  New G-protein-coupled receptor crystal structures: insights and limitations. , 2008, Trends in pharmacological sciences.

[32]  Stefano Costanzi,et al.  Computing Highly Correlated Positions Using Mutual Information and Graph Theory for G Protein-Coupled Receptors , 2009, PloS one.

[33]  G Vriend,et al.  Correlated Mutation Analyses on Very Large Sequence Families , 2002, Chembiochem : a European journal of chemical biology.

[34]  Robert D. Finn,et al.  Pfam: clans, web tools and services , 2005, Nucleic Acids Res..

[35]  J. Hebebrand,et al.  A Heterozygous Mutation in the Third Transmembrane Domain Causes a Dominant-Negative Effect on Signalling Capability of the MC4R , 2008, Obesity Facts.

[36]  Gerhard Hessler,et al.  Drug Design Strategies for Targeting G‐Protein‐Coupled Receptors , 2002, Chembiochem : a European journal of chemical biology.

[37]  J. Wess,et al.  Molecular basis of receptor/G-protein-coupling selectivity. , 1998, Pharmacology & therapeutics.

[38]  Martin Jones,et al.  IUPHAR-DB: the IUPHAR database of G protein-coupled receptors and ion channels , 2008, Nucleic Acids Res..

[39]  Kurt Kristiansen,et al.  Molecular mechanisms of ligand binding, signaling, and regulation within the superfamily of G-protein-coupled receptors: molecular modeling and mutagenesis approaches to receptor structure and function. , 2004, Pharmacology & therapeutics.

[40]  Stephen K.-F. Wong,et al.  G Protein Selectivity Is Regulated by Multiple Intracellular Regions of GPCRs , 2003, Neurosignals.

[41]  U. Zabel,et al.  Fluorescence Resonance Energy Transfer Analysis of α2a-Adrenergic Receptor Activation Reveals Distinct Agonist-Specific Conformational Changes , 2009, Molecular Pharmacology.

[42]  R. Paschke,et al.  Principles and Determinants of G-Protein Coupling by the Rhodopsin-Like Thyrotropin Receptor , 2010, PloS one.

[43]  Lemont B. Kier,et al.  The E-State as the Basis for Molecular Structure Space Definition and Structure Similarity , 2000, J. Chem. Inf. Comput. Sci..

[44]  Didier Rognan,et al.  A chemogenomic analysis of the transmembrane binding cavity of human G‐protein‐coupled receptors , 2005, Proteins.

[45]  Robert P Bywater,et al.  Location and nature of the residues important for ligand recognition in G‐protein coupled receptors , 2005, Journal of molecular recognition : JMR.

[46]  F. Ozoe,et al.  Single amino acid of an octopamine receptor as a molecular switch for distinct G protein couplings. , 2008, Biochemical and biophysical research communications.

[47]  G. Vauquelin,et al.  G protein‐coupled receptors: a count of 1001 conformations , 2005, Fundamental & clinical pharmacology.

[48]  G. Krause,et al.  Thyrotropin and homologous glycoprotein hormone receptors: structural and functional aspects of extracellular signaling mechanisms. , 2009, Endocrine reviews.

[49]  D. Gloriam,et al.  Definition of the G protein-coupled receptor transmembrane bundle binding pocket and calculation of receptor similarities for drug design. , 2009, Journal of medicinal chemistry.

[50]  Catherine L. Worth,et al.  Comparative Sequence and Structural Analyses of G-Protein-Coupled Receptor Crystal Structures and Implications for Molecular Models , 2009, PloS one.

[51]  T. Schöneberg,et al.  Learning from the past: evolution of GPCR functions. , 2007, Trends in pharmacological sciences.

[52]  Sadashiva S Karnik,et al.  Multiple Signaling States of G-Protein-Coupled Receptors , 2005, Pharmacological Reviews.

[53]  Jan Kelder,et al.  A signaling-selective, nanomolar potent allosteric low molecular weight agonist for the human luteinizing hormone receptor , 2008, Naunyn-Schmiedeberg's Archives of Pharmacology.

[54]  R. Stevens,et al.  High-Resolution Crystal Structure of an Engineered Human β2-Adrenergic G Protein–Coupled Receptor , 2007, Science.

[55]  M. Mulholland,et al.  Interactions of human melanocortin 4 receptor with nonpeptide and peptide agonists. , 2005, Biochemistry.

[56]  J. Wess,et al.  Hydrophilic side chains in the third and seventh transmembrane helical domains of human A2A adenosine receptors are required for ligand recognition. , 1996, Molecular pharmacology.

[57]  H. Hamm,et al.  Heterotrimeric G protein activation by G-protein-coupled receptors , 2008, Nature Reviews Molecular Cell Biology.

[58]  Oliver P. Ernst,et al.  A Ligand Channel through the G Protein Coupled Receptor Opsin , 2009, PloS one.

[59]  Rob Leurs,et al.  Pharmacogenomic and structural analysis of constitutive g protein-coupled receptor activity. , 2007, Annual review of pharmacology and toxicology.

[60]  A. Smrcka.  G protein βγ subunits: Central mediators of G protein-coupled receptor signaling , 2008, Cellular and Molecular Life Sciences.

[61]  K. Wanner,et al.  Methods and Principles in Medicinal Chemistry , 2007 .

[62]  Jianping Lin,et al.  Allosteric antagonist binding sites in class B GPCRs: corticotropin receptor 1 , 2010, J. Comput. Aided Mol. Des..

[63]  Xavier Deupi,et al.  Conformational complexity of G-protein-coupled receptors. , 2007, Trends in pharmacological sciences.

[64]  Michael J. Keiser,et al.  Relating protein pharmacology by ligand chemistry , 2007, Nature Biotechnology.

[65]  J. Wess,et al.  Structural basis of G protein-coupled receptor-G protein interactions. , 2010, Nature chemical biology.

[66]  Vsevolod Katritch,et al.  Ligand binding and subtype selectivity of the human A(2A) adenosine receptor: identification and characterization of essential amino acid residues. , 2010, The Journal of biological chemistry.

[67]  S. Karnik,et al.  Ligand-specific Conformation of Extracellular Loop-2 in the Angiotensin II Type 1 Receptor* , 2010, The Journal of Biological Chemistry.

[68]  S. Rasmussen,et al.  The structure and function of G-protein-coupled receptors , 2009, Nature.

[69]  R. Seifert,et al.  Constitutive activity of G-protein-coupled receptors: cause of disease and common property of wild-type receptors , 2002, Naunyn-Schmiedeberg's Archives of Pharmacology.

[70]  R. Seifert,et al.  Molecular analysis of beta(2)-adrenoceptor coupling to G(s)-, G(i)-, and G(q)-proteins. , 2000, Molecular pharmacology.

[71]  K. Fahmy,et al.  Linkage between the intramembrane H-bond network around aspartic acid 83 and the cytosolic environment of helix 8 in photoactivated rhodopsin. , 2007, Journal of molecular biology.

[72]  Leonardo Pardo,et al.  An Activation Switch in the Rhodopsin Family of G Protein-coupled Receptors , 2005, Journal of Biological Chemistry.

[73]  J. Ballesteros,et al.  [19] Integrated methods for the construction of three-dimensional models and computational probing of structure-function relations in G protein-coupled receptors , 1995 .

[74]  Katrin Sangkuhl,et al.  Mutant G-protein-coupled receptors as a cause of human diseases. , 2004, Pharmacology & therapeutics.

[75]  Roberto Todeschini,et al.  Handbook of Molecular Descriptors , 2002 .

[76]  P. Evans,et al.  Agonist-specific coupling of G-protein-coupled receptors to second-messenger systems. , 1995, Progress in brain research.

[77]  A. Ghose,et al.  Prediction of Hydrophobic (Lipophilic) Properties of Small Organic Molecules Using Fragmental Methods: An Analysis of ALOGP and CLOGP Methods , 1998 .

[78]  Oliver P. Ernst,et al.  Crystal structure of opsin in its G-protein-interacting conformation , 2008, Nature.

[79]  P. Scheerer,et al.  A G protein-coupled receptor at work: the rhodopsin model. , 2009, Trends in biochemical sciences.

[80]  G. Schertler.  Signal transduction: The rhodopsin story continued , 2008, Nature.

[81]  Lemont B. Kier,et al.  The electrotopological state: structure information at the atomic level for molecular graphs , 1991, J. Chem. Inf. Comput. Sci..

[82]  Jaak Vilo,et al.  Prediction of the coupling specificity of G protein coupled receptors to their G proteins , 2001, ISMB.

[83]  Carsten O. Daub,et al.  The mutual information: Detecting and evaluating dependencies between variables , 2002, ECCB.

[84]  R. Horuk,et al.  I want a new drug: G-protein-coupled receptors in drug development. , 2006, Drug discovery today.

[85]  M. Maccoss,et al.  Characterization of a Novel, Non-peptidyl Antagonist of the Human Glucagon Receptor* , 1999, The Journal of Biological Chemistry.

[86]  L. Pardo,et al.  Helix 8 of the Viral Chemokine Receptor ORF74 Directs Chemokine Binding* , 2006, Journal of Biological Chemistry.

[87]  Xin-Yun Huang,et al.  When a G protein-coupled receptor does not couple to a G protein. , 2007, Molecular bioSystems.

[88]  Hugo Kubinyi,et al.  Chemogenomics in Drug Discovery: A Medicinal Chemistry Perspective , 2004 .

[89]  Raymond C Stevens,et al.  Discovery of new GPCR biology: one receptor structure at a time. , 2009, Structure.

[90]  R. Maki,et al.  Molecular Determinants of Melanocortin 4 Receptor Ligand Binding and MC4/MC3 Receptor Selectivity , 2003, Journal of Pharmacology and Experimental Therapeutics.

[91]  A. Hopkins,et al.  The druggable genome , 2002, Nature Reviews Drug Discovery.

[92]  J. Wess,et al.  Conformational changes involved in G-protein-coupled-receptor activation. , 2008, Trends in pharmacological sciences.

[93]  Sean R. Eddy,et al.  Profile hidden Markov models , 1998, Bioinform..

[94]  M. Rooman,et al.  Evidence that Interaction between Conserved Residues in Transmembrane Helices 2, 3, and 7 Are Crucial for Human VPAC1 Receptor Activation , 2010, Molecular Pharmacology.

[95]  Kai Ye,et al.  A two‐entropies analysis to identify functional positions in the transmembrane region of class A G protein‐coupled receptors , 2006, Proteins.