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PMID: 25589408 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Review

Methodological advances: the unsung heroes of the GPCR structural revolution.

Nature reviews. Molecular cell biology ·Vol. 16 ·No. 2 ·2015-02-00 ·Pages 69-81

Ghosh E, Kumari P, Jaiman D, Shukla AK

Abstract

G protein-coupled receptors (GPCRs) are intricately involved in a diverse array of physiological processes and pathophysiological conditions. They constitute the largest class of drug target in the human genome, which highlights the importance of understanding the molecular basis of their activation, downstream signalling and regulation. In the past few years, considerable progress has been made in our ability to visualize GPCRs and their signalling complexes at the structural level. This is due to a series of methodological developments, improvements in technology and the use of highly innovative approaches, such as protein engineering, new detergents, lipidic cubic phase-based crystallization and microfocus synchrotron beamlines. These advances suggest that an unprecedented amount of structural information will become available in the field of GPCR biology in the coming years.

MeSH Terms
Animals Humans Protein Conformation Protein Engineering/methods Receptors, G-Protein-Coupled/chemistry,genetics Signal Transduction/genetics
Chemicals
Receptors, G-Protein-Coupled
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Ghosh Eshan
Department of Biological Sciences and Bioengineering, Indian Institute of Technology, Kanpur 208016, India.
Kumari Punita
Department of Biological Sciences and Bioengineering, Indian Institute of Technology, Kanpur 208016, India.
Jaiman Deepika
Department of Biological Sciences and Bioengineering, Indian Institute of Technology, Kanpur 208016, India.
Shukla Arun K
Department of Biological Sciences and Bioengineering, Indian Institute of Technology, Kanpur 208016, India.
References (120)
120 references, click to expand
  1. Structure of class C GPCR metabotropic glutamate receptor 5 transmembrane domain.
    Nature. 2014 Jul 31;511(7511):557-62 PMID: 25042998
  2. G-protein-coupled receptor inactivation by an allosteric inverse-agonist antibody.
    Nature. 2012 Jan 29;482(7384):237-40 PMID: 22286059
  3. G protein-coupled receptor trafficking in health and disease: lessons learned to prepare for therapeutic mutant rescue in vivo.
    Pharmacol Rev. 2007 Sep;59(3):225-50 PMID: 17878512
  4. β-arrestins and G protein-coupled receptor trafficking.
    Handb Exp Pharmacol. 2014;219:173-86 PMID: 24292830
  5. Structure and function of an irreversible agonist-β(2) adrenoceptor complex.
    Nature. 2011 Jan 13;469(7329):236-40 PMID: 21228876
  6. International Union of Pharmacology. XLVI. G protein-coupled receptor list.
    Pharmacol Rev. 2005 Jun;57(2):279-88 PMID: 15914470
  7. Structure of the human histamine H1 receptor complex with doxepin.
    Nature. 2011 Jun 22;475(7354):65-70 PMID: 21697825
  8. Structure of the human κ-opioid receptor in complex with JDTic.
    Nature. 2012 Mar 21;485(7398):327-32 PMID: 22437504
  9. A crystal clear view of the beta2-adrenergic receptor.
    Nat Biotechnol. 2008 Feb;26(2):189-91 PMID: 18259173
  10. N-terminal T4 lysozyme fusion facilitates crystallization of a G protein coupled receptor.
    PLoS One. 2012;7(10):e46039 PMID: 23056231
  11. Production and characterization of monoclonal antibodies directed against native epitopes of NhaA, the Na+/H+ antiporter of Escherichia coli.
    FEBS Lett. 1998 Dec 11;441(1):53-8 PMID: 9877164
  12. Structural flexibility of the G alpha s alpha-helical domain in the beta2-adrenoceptor Gs complex.
    Proc Natl Acad Sci U S A. 2011 Sep 20;108(38):16086-91 PMID: 21914848
  13. Affinity chromatography of the beta-adrenergic receptor.
    J Biol Chem. 1979 Apr 25;254(8):2923-7 PMID: 218957
  14. Seven-transmembrane receptors.
    Nat Rev Mol Cell Biol. 2002 Sep;3(9):639-50 PMID: 12209124
  15. Crystal structure of the ligand-free G-protein-coupled receptor opsin.
    Nature. 2008 Jul 10;454(7201):183-7 PMID: 18563085
  16. Crucial steps in the structure determination of the Na+/H+ antiporter NhaA in its native conformation.
    J Mol Biol. 2006 Sep 15;362(2):192-202 PMID: 16919297
  17. Structural basis for allosteric regulation of GPCRs by sodium ions.
    Science. 2012 Jul 13;337(6091):232-6 PMID: 22798613
  18. Agonists induce conformational changes in transmembrane domains III and VI of the beta2 adrenoceptor.
    EMBO J. 1997 Nov 17;16(22):6737-47 PMID: 9362488
  19. Comparative analysis of the human angiotensin II type 1a receptor heterologously produced in insect cells and mammalian cells.
    Biochem Biophys Res Commun. 2006 Oct 13;349(1):6-14 PMID: 16963356
  20. A general protocol for the generation of Nanobodies for structural biology.
    Nat Protoc. 2014 Mar;9(3):674-93 PMID: 24577359
  21. Lipidic cubic phase injector facilitates membrane protein serial femtosecond crystallography.
    Nat Commun. 2014;5:3309 PMID: 24525480
  22. Structure of the human glucagon class B G-protein-coupled receptor.
    Nature. 2013 Jul 25;499(7459):444-9 PMID: 23863937
  23. GRKs and beta-arrestins: roles in receptor silencing, trafficking and signaling.
    Trends Endocrinol Metab. 2006 May-Jun;17(4):159-65 PMID: 16595179
  24. X-ray structure of bacteriorhodopsin at 2.5 angstroms from microcrystals grown in lipidic cubic phases.
    Science. 1997 Sep 12;277(5332):1676-81 PMID: 9287223
  25. Preparation of microcrystals in lipidic cubic phase for serial femtosecond crystallography.
    Nat Protoc. 2014 Sep;9(9):2123-34 PMID: 25122522
  26. Crystallizing membrane proteins using lipidic mesophases.
    Nat Protoc. 2009;4(5):706-31 PMID: 19390528
  27. Biochemical and pharmacological characterization of the human bradykinin subtype 2 receptor produced in mammalian cells using the Semliki Forest virus system.
    Biol Chem. 2006 May;387(5):569-76 PMID: 16740128
  28. LCP-FRAP Assay for Pre-Screening Membrane Proteins for in Meso Crystallization.
    Cryst Growth Des. 2008;8(12):4307-4315 PMID: 19234616
  29. Structure of signaling-competent neurotensin receptor 1 obtained by directed evolution in Escherichia coli.
    Proc Natl Acad Sci U S A. 2014 Feb 11;111(6):E655-62 PMID: 24453215
  30. Structures of the CXCR4 chemokine GPCR with small-molecule and cyclic peptide antagonists.
    Science. 2010 Nov 19;330(6007):1066-71 PMID: 20929726
  31. Beta-arrestin acts as a clathrin adaptor in endocytosis of the beta2-adrenergic receptor.
    Nature. 1996 Oct 3;383(6599):447-50 PMID: 8837779
  32. Amino and carboxyl terminal modifications to facilitate the production and purification of a G protein-coupled receptor.
    Anal Biochem. 1995 Oct 10;231(1):269-71 PMID: 8678314
  33. Covalent agonists for studying G protein-coupled receptor activation.
    Proc Natl Acad Sci U S A. 2014 Jul 22;111(29):10744-8 PMID: 25006259
  34. Crystal structure of the human beta2 adrenergic G-protein-coupled receptor.
    Nature. 2007 Nov 15;450(7168):383-7 PMID: 17952055
  35. Fusion partner toolchest for the stabilization and crystallization of G protein-coupled receptors.
    Structure. 2012 Jun 6;20(6):967-76 PMID: 22681902
  36. Structure of class B GPCR corticotropin-releasing factor receptor 1.
    Nature. 2013 Jul 25;499(7459):438-43 PMID: 23863939
  37. Visualization of arrestin recruitment by a G-protein-coupled receptor.
    Nature. 2014 Aug 14;512(7513):218-222 PMID: 25043026
  38. Crystal structure of a common GPCR-binding interface for G protein and arrestin.
    Nat Commun. 2014 Sep 10;5:4801 PMID: 25205354
  39. Molecular mechanism for the crystallization of bacteriorhodopsin in lipidic cubic phases.
    FEBS Lett. 2001 Aug 31;504(3):179-86 PMID: 11532451
  40. Lipidic Cubic Phases: New Matrices for the Three-Dimensional Crystallization of Membrane Proteins.
    J Struct Biol. 1998;121(2):82-91 PMID: 9618339
  41. Molecular mechanism of β-arrestin-biased agonism at seven-transmembrane receptors.
    Annu Rev Pharmacol Toxicol. 2012;52:179-97 PMID: 21942629
  42. Role of β-arrestins and arrestin domain-containing proteins in G protein-coupled receptor trafficking.
    Curr Opin Cell Biol. 2014 Apr;27:63-71 PMID: 24680432
  43. Conformational changes in the G protein Gs induced by the β2 adrenergic receptor.
    Nature. 2011 Sep 28;477(7366):611-5 PMID: 21956331
  44. Crystal structure of the µ-opioid receptor bound to a morphinan antagonist.
    Nature. 2012 Mar 21;485(7398):321-6 PMID: 22437502
  45. Heptahelical receptor signaling: beyond the G protein paradigm.
    J Cell Biol. 1999 May 31;145(5):927-32 PMID: 10352011
  46. Fv fragment-mediated crystallization of the membrane protein bacterial cytochrome c oxidase.
    Nat Struct Biol. 1995 Oct;2(10):842-6 PMID: 7552705
  47. Structure of the CCR5 chemokine receptor-HIV entry inhibitor maraviroc complex.
    Science. 2013 Sep 20;341(6152):1387-90 PMID: 24030490
  48. Study of a synthetic human olfactory receptor 17-4: expression and purification from an inducible mammalian cell line.
    PLoS One. 2008 Aug 06;3(8):e2920 PMID: 18682799
  49. Crystal structure of the β2 adrenergic receptor-Gs protein complex.
    Nature. 2011 Jul 19;477(7366):549-55 PMID: 21772288
  50. The 2.6 angstrom crystal structure of a human A2A adenosine receptor bound to an antagonist.
    Science. 2008 Nov 21;322(5905):1211-7 PMID: 18832607
  51. G protein-coupled receptors: abnormalities in signal transmission, disease states and pharmacotherapy.
    Acta Pol Pharm. 2014 Mar-Apr;71(2):229-43 PMID: 25272642
  52. The synthesis and high-level expression of a beta2-adrenergic receptor gene in a tetracycline-inducible stable mammalian cell line.
    Protein Sci. 2006 Jun;15(6):1433-40 PMID: 16731977
  53. Crystal structure of a thermally stable rhodopsin mutant.
    J Mol Biol. 2007 Oct 5;372(5):1179-88 PMID: 17825322
  54. Structure of the δ-opioid receptor bound to naltrindole.
    Nature. 2012 May 16;485(7398):400-4 PMID: 22596164
  55. Value of novelty?
    Nat Rev Drug Discov. 2002 Aug;1(8):571-2 PMID: 12402497
  56. Engineering G protein-coupled receptors to facilitate their structure determination.
    Curr Opin Struct Biol. 2009 Aug;19(4):386-95 PMID: 19682887
  57. Activation and allosteric modulation of a muscarinic acetylcholine receptor.
    Nature. 2013 Dec 5;504(7478):101-6 PMID: 24256733
  58. Synthesis and properties of dodecyl trehaloside detergents for membrane protein studies.
    Langmuir. 2012 Jul 31;28(30):11173-81 PMID: 22780816
  59. Functional overexpression and characterization of human bradykinin subtype 2 receptor in insect cells using the baculovirus system.
    J Cell Biochem. 2006 Oct 15;99(3):868-77 PMID: 16721823
  60. Structure and dynamics of the M3 muscarinic acetylcholine receptor.
    Nature. 2012 Feb 22;482(7386):552-6 PMID: 22358844
  61. Structure of active β-arrestin-1 bound to a G-protein-coupled receptor phosphopeptide.
    Nature. 2013 May 2;497(7447):137-41 PMID: 23604254
  62. Use of fluorescence spectroscopy to study conformational changes in the beta 2-adrenoceptor.
    Methods Enzymol. 2002;343:170-82 PMID: 11665566
  63. Distinct conformational changes in beta-arrestin report biased agonism at seven-transmembrane receptors.
    Proc Natl Acad Sci U S A. 2008 Jul 22;105(29):9988-93 PMID: 18621717
  64. High-resolution structure of the human GPR40 receptor bound to allosteric agonist TAK-875.
    Nature. 2014 Sep 4;513(7516):124-7 PMID: 25043059
  65. Conserved activation pathways in G-protein-coupled receptors.
    Biochem Soc Trans. 2012 Apr;40(2):383-8 PMID: 22435816
  66. High-resolution crystal structure of an engineered human beta2-adrenergic G protein-coupled receptor.
    Science. 2007 Nov 23;318(5854):1258-65 PMID: 17962520
  67. Employing Rhodobacter sphaeroides to functionally express and purify human G protein-coupled receptors.
    Biol Chem. 2008 Jan;389(1):69-78 PMID: 18095871
  68. Structure of the human dopamine D3 receptor in complex with a D2/D3 selective antagonist.
    Science. 2010 Nov 19;330(6007):1091-5 PMID: 21097933
  69. Structure of the nociceptin/orphanin FQ receptor in complex with a peptide mimetic.
    Nature. 2012 May 16;485(7398):395-9 PMID: 22596163
  70. Crystal structure of oligomeric β1-adrenergic G protein-coupled receptors in ligand-free basal state.
    Nat Struct Mol Biol. 2013 Apr;20(4):419-25 PMID: 23435379
  71. Structure of a nanobody-stabilized active state of the β(2) adrenoceptor.
    Nature. 2011 Jan 13;469(7329):175-80 PMID: 21228869
  72. Emerging paradigms of β-arrestin-dependent seven transmembrane receptor signaling.
    Trends Biochem Sci. 2011 Sep;36(9):457-69 PMID: 21764321
  73. GPCR engineering yields high-resolution structural insights into beta2-adrenergic receptor function.
    Science. 2007 Nov 23;318(5854):1266-73 PMID: 17962519
  74. Crystal structure of a lipid G protein-coupled receptor.
    Science. 2012 Feb 17;335(6070):851-5 PMID: 22344443
  75. Structure of the agonist-bound neurotensin receptor.
    Nature. 2012 Oct 25;490(7421):508-13 PMID: 23051748
  76. A specific cholesterol binding site is established by the 2.8 A structure of the human beta2-adrenergic receptor.
    Structure. 2008 Jun;16(6):897-905 PMID: 18547522
  77. Serial femtosecond crystallography of G protein-coupled receptors.
    Science. 2013 Dec 20;342(6165):1521-4 PMID: 24357322
  78. Production of a bioengineered G-protein coupled receptor of human formyl peptide receptor 3.
    PLoS One. 2011;6(8):e23076 PMID: 21853070
  79. The structural basis of arrestin-mediated regulation of G-protein-coupled receptors.
    Pharmacol Ther. 2006 Jun;110(3):465-502 PMID: 16460808
  80. Glucose-neopentyl glycol (GNG) amphiphiles for membrane protein study.
    Chem Commun (Camb). 2013 Mar 21;49(23):2287-9 PMID: 23165475
  81. Biasing GPCR signaling from inside.
    Sci Signal. 2014 Jan 28;7(310):pe3 PMID: 24473194
  82. Femtosecond X-ray protein nanocrystallography.
    Nature. 2011 Feb 3;470(7332):73-7 PMID: 21293373
  83. LCP-Tm: an assay to measure and understand stability of membrane proteins in a membrane environment.
    Biophys J. 2010 Apr 21;98(8):1539-48 PMID: 20409473
  84. Maltose-neopentyl glycol (MNG) amphiphiles for solubilization, stabilization and crystallization of membrane proteins.
    Nat Methods. 2010 Dec;7(12):1003-8 PMID: 21037590
  85. Structural determinants of arrestin functions.
    Prog Mol Biol Transl Sci. 2013;118:57-92 PMID: 23764050
  86. Room to move: crystallizing membrane proteins in swollen lipidic mesophases.
    J Mol Biol. 2006 Apr 14;357(5):1605-18 PMID: 16490208
  87. Seven transmembrane receptors: something old, something new.
    Acta Physiol (Oxf). 2007 May;190(1):9-19 PMID: 17428228
  88. Molecular tinkering of G protein-coupled receptors: an evolutionary success.
    EMBO J. 1999 Apr 1;18(7):1723-9 PMID: 10202136
  89. Crystallization of bacteriorhodopsin from bicelle formulations at room temperature.
    Protein Sci. 2005 Mar;14(3):836-40 PMID: 15689517
  90. Ligand-specific regulation of the extracellular surface of a G-protein-coupled receptor.
    Nature. 2010 Jan 7;463(7277):108-12 PMID: 20054398
  91. Structure of a beta1-adrenergic G-protein-coupled receptor.
    Nature. 2008 Jul 24;454(7203):486-91 PMID: 18594507
  92. Nanobody stabilization of G protein-coupled receptor conformational states.
    Curr Opin Struct Biol. 2011 Aug;21(4):567-72 PMID: 21782416
  93. GPCR stabilization using the bicelle-like architecture of mixed sterol-detergent micelles.
    Methods. 2011 Dec;55(4):310-7 PMID: 22041719
  94. A robotic system for crystallizing membrane and soluble proteins in lipidic mesophases.
    Acta Crystallogr D Biol Crystallogr. 2004 Oct;60(Pt 10):1795-807 PMID: 15388926
  95. Heterologous expression and characterization of the recombinant bradykinin B2 receptor using the methylotrophic yeast Pichia pastoris.
    Protein Expr Purif. 2007 Sep;55(1):1-8 PMID: 17711791
  96. Crystallization in cubo: general applicability to membrane proteins.
    Acta Crystallogr D Biol Crystallogr. 2000 Jun;56(Pt 6):781-4 PMID: 10818364
  97. Development and crystallization of a minimal thermostabilised G protein-coupled receptor.
    Protein Expr Purif. 2009 Jun;65(2):204-13 PMID: 19297694
  98. The molecular acrobatics of arrestin activation.
    Trends Pharmacol Sci. 2004 Feb;25(2):105-11 PMID: 15102497
  99. Crystallisation of membrane proteins mediated by antibody fragments.
    Curr Opin Struct Biol. 2002 Aug;12(4):503-8 PMID: 12163074
  100. Structure at 2.3 A resolution of the cytochrome bc(1) complex from the yeast Saccharomyces cerevisiae co-crystallized with an antibody Fv fragment.
    Structure. 2000 Jun 15;8(6):669-84 PMID: 10873857
  101. Structural genomics on membrane proteins: comparison of more than 100 GPCRs in 3 expression systems.
    J Struct Funct Genomics. 2006 Jun;7(2):77-91 PMID: 17120110
  102. Beta-arrestins and cell signaling.
    Annu Rev Physiol. 2007;69:483-510 PMID: 17305471
  103. Novel tripod amphiphiles for membrane protein analysis.
    Chemistry. 2013 Nov 11;19(46):15645-51 PMID: 24123610
  104. High-resolution crystal structure of human protease-activated receptor 1.
    Nature. 2012 Dec 20;492(7429):387-92 PMID: 23222541
  105. Enhancing functional production of G protein-coupled receptors in Pichia pastoris to levels required for structural studies via a single expression screen.
    Protein Sci. 2006 May;15(5):1115-26 PMID: 16597836
  106. Lipidic cubic phases: a novel concept for the crystallization of membrane proteins.
    Proc Natl Acad Sci U S A. 1996 Dec 10;93(25):14532-5 PMID: 8962086
  107. Bicelle crystallization: a new method for crystallizing membrane proteins yields a monomeric bacteriorhodopsin structure.
    J Mol Biol. 2002 Feb 8;316(1):1-6 PMID: 11829498
  108. Crystallizing thinking about the beta2-adrenergic receptor.
    Mol Pharmacol. 2008 May;73(5):1333-8 PMID: 18239031
  109. Dimethylsulphoxide as a tool to increase functional expression of heterologously produced GPCRs in mammalian cells.
    FEBS Lett. 2006 Jul 24;580(17):4261-5 PMID: 16831432
  110. Steroid-based facial amphiphiles for stabilization and crystallization of membrane proteins.
    Proc Natl Acad Sci U S A. 2013 Mar 26;110(13):E1203-11 PMID: 23479627
  111. Heterologous expression and comparative characterization of the human neuromedin U subtype II receptor using the methylotrophic yeast Pichia pastoris and mammalian cells.
    Int J Biochem Cell Biol. 2007;39(5):931-42 PMID: 17445746
  112. Structure of the human M2 muscarinic acetylcholine receptor bound to an antagonist.
    Nature. 2012 Jan 25;482(7386):547-51 PMID: 22278061
  113. Adrenaline-activated structure of β2-adrenoceptor stabilized by an engineered nanobody.
    Nature. 2013 Oct 24;502(7472):575-579 PMID: 24056936
  114. Structural diversity of G protein-coupled receptors and significance for drug discovery.
    Nat Rev Drug Discov. 2008 Apr;7(4):339-57 PMID: 18382464
  115. Structure-function of the G protein-coupled receptor superfamily.
    Annu Rev Pharmacol Toxicol. 2013;53:531-56 PMID: 23140243
  116. Membrane protein crystallization in lipidic mesophases with tailored bilayers.
    Structure. 2004 Dec;12(12):2113-24 PMID: 15576026
  117. Structure of the adenosine A(2A) receptor in complex with ZM241385 and the xanthines XAC and caffeine.
    Structure. 2011 Sep 7;19(9):1283-93 PMID: 21885291
  118. Rastering strategy for screening and centring of microcrystal samples of human membrane proteins with a sub-10 microm size X-ray synchrotron beam.
    J R Soc Interface. 2009 Oct 6;6 Suppl 5:S587-97 PMID: 19535414
  119. Development of an Automated High Throughput LCP-FRAP Assay to Guide Membrane Protein Crystallization in Lipid Mesophases.
    Cryst Growth Des. 2011 Apr 6;11(4):1193-1201 PMID: 21660116
  120. Coupling ligand structure to specific conformational switches in the beta2-adrenoceptor.
    Nat Chem Biol. 2006 Aug;2(8):417-22 PMID: 16799554
Article Info
Journal
Nature reviews. Molecular cell biology
Abbr.
Nat Rev Mol Cell Biol
ISSN
1471-0080
DOI
10.1038/nrm3933
Published
2015-02-00
Epub
2015-00-15
Pages
69-81
Language
English
Region
England
NLM ID
100962782
Subset
IM
External Links
PubMed source DOI: 10.1038/nrm3933 View journal details
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