Current applications of mini G proteins to study the structure and function of G protein-coupled receptors

Byron Carpenter; Byron Carpenter

doi:10.3934/bioeng.2018.4.209

AIMS Bioengineering

2018, Volume 5, Issue 4: 209-225. doi: 10.3934/bioeng.2018.4.209

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Review

Current applications of mini G proteins to study the structure and function of G protein-coupled receptors

Byron Carpenter ^,

Warwick Integrative Synthetic Biology (WISB) centre, School of Life Sciences, University of Warwick, Coventry CV4 7AL, UK

Received: 29 August 2018 Accepted: 12 October 2018 Published: 17 October 2018

G protein-coupled receptors (GPCRs) regulate intracellular signalling pathways that contribute to virtually all aspects of cell function. Characterising GPCRs in each of their conformational states is key to understanding their mechanism of action, but structure determination of receptors in their active state, bound to a heterotrimeric G protein or b-arrestin, has proved challenging. A number of G protein surrogates have been developed to simplify this process, including G protein-derived peptides, nanobodies and, most recently, mini G proteins. The aim of these surrogates is to bind the receptor and stabilise its active conformation, whilst eliminating the problems inherent to native signalling proteins, namely their large size, instability and conformational dynamics. Mini G proteins are composed of a single domain from the G protein a-subunit that has been engineered to form a stable complex with GPCRs. They induce comparable pharmacological and structural changes in the receptor to those elicited by heterotrimeric G proteins, and retain their native receptor-coupling specificity. At least one member of each G protein family has been converted into a mini G protein, which means that they can be used to characterise a wide variety of GPCRs. Since their initial publication two years ago, mini G proteins have facilitated the structure determination of three different receptors in their active state and enabled the development of a methodology to thermostabilise GPCRs in their fully active conformation. They have also been used to develop a range of assays that can measure mini G protein coupling to receptors in vitro, and a sensitive cell-based assay that is capable of accurately reporting ligand efficacy and quantifying G protein coupling in vivo. This review presents an overview of the current applications of mini G proteins to study the structure and function of GPCRs.
- active state,
- GPCR,
- G protein-coupled receptor,
- heterotrimeric G protein,
- mini G protein,
- mini-Gs,
- signalling complex
Citation: Byron Carpenter. Current applications of mini G proteins to study the structure and function of G protein-coupled receptors[J]. AIMS Bioengineering, 2018, 5(4): 209-225. doi: 10.3934/bioeng.2018.4.209

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Abstract

G protein-coupled receptors (GPCRs) regulate intracellular signalling pathways that contribute to virtually all aspects of cell function. Characterising GPCRs in each of their conformational states is key to understanding their mechanism of action, but structure determination of receptors in their active state, bound to a heterotrimeric G protein or b-arrestin, has proved challenging. A number of G protein surrogates have been developed to simplify this process, including G protein-derived peptides, nanobodies and, most recently, mini G proteins. The aim of these surrogates is to bind the receptor and stabilise its active conformation, whilst eliminating the problems inherent to native signalling proteins, namely their large size, instability and conformational dynamics. Mini G proteins are composed of a single domain from the G protein a-subunit that has been engineered to form a stable complex with GPCRs. They induce comparable pharmacological and structural changes in the receptor to those elicited by heterotrimeric G proteins, and retain their native receptor-coupling specificity. At least one member of each G protein family has been converted into a mini G protein, which means that they can be used to characterise a wide variety of GPCRs. Since their initial publication two years ago, mini G proteins have facilitated the structure determination of three different receptors in their active state and enabled the development of a methodology to thermostabilise GPCRs in their fully active conformation. They have also been used to develop a range of assays that can measure mini G protein coupling to receptors in vitro, and a sensitive cell-based assay that is capable of accurately reporting ligand efficacy and quantifying G protein coupling in vivo. This review presents an overview of the current applications of mini G proteins to study the structure and function of GPCRs.

References

[1]	Wise A, Gearing K, Rees S (2002) Target validation of G-protein coupled receptors. Drug Discov Today 7: 235–246. doi: 10.1016/S1359-6446(01)02131-6
[2]	Cherezov V, Rosenbaum DM, Hanson MA, et al. (2007) High-resolution crystal structure of an engineered human beta2-adrenergic G protein-coupled receptor. Science 318: 1258–1265. doi: 10.1126/science.1150577
[3]	Serrano-Vega MJ, Magnani F, Shibata Y, et al. (2008) Conformational thermostabilization of the beta1-adrenergic receptor in a detergent-resistant form. Proc Natl Acad Sci USA 105: 877–882. doi: 10.1073/pnas.0711253105
[4]	Caffrey M (2015) A comprehensive review of the lipid cubic phase or in meso method for crystallizing membrane and soluble proteins and complexes. Acta Crystallogr F Struct Biol Commun 71: 3–18.
[5]	Landau EM, Rosenbusch JP (1996) Lipidic cubic phases: A novel concept for the crystallization of membrane proteins. Proc Natl Acad Sci USA 93: 14532–14535. doi: 10.1073/pnas.93.25.14532
[6]	Chan W, Said M, Zhang C, et al. (2018) GPCR-EXP: A semi-manually curated database for experimentally-solved and predicted GPCR structures. Available from: https://zhanglab.ccmb.med.umich.edu/GPCR-EXP/.
[7]	Jazayeri A, Dias JM, Marshall FH (2015) From G Protein-coupled receptor structure resolution to rational drug design. J Biol Chem 290: 19489–19495. doi: 10.1074/jbc.R115.668251
[8]	Carpenter B, Tate CG (2017) Active state structures of G protein-coupled receptors highlight the similarities and differences in the G protein and arrestin coupling interfaces. Curr Opin Struct Biol 45: 124–132. doi: 10.1016/j.sbi.2017.04.010
[9]	Rasmussen SG, Devree BT, Zou Y, et al. (2011) Crystal structure of the beta2 adrenergic receptor-Gs protein complex. Nature 477: 549–555. doi: 10.1038/nature10361
[10]	Kang Y, Zhou XE, Gao X, et al. (2015) Crystal structure of rhodopsin bound to arrestin by femtosecond X-ray laser. Nature 523: 561–567. doi: 10.1038/nature14656
[11]	Westfield GH, Rasmussen SG, Su M, et al. (2011) Structural flexibility of the G alpha s alpha-helical domain in the beta2-adrenoceptor Gs complex. Proc Natl Acad Sci USA 108: 16086–16091. doi: 10.1073/pnas.1113645108
[12]	Van EN, Preininger AM, Alexander N, et al. (2011) Interaction of a G protein with an activated receptor opens the interdomain interface in the alpha subunit. Proc Natl Acad Sci USA 108: 9420–9424. doi: 10.1073/pnas.1105810108
[13]	Scheerer P, Park JH, Hildebrand PW, et al. (2008) Crystal structure of opsin in its G-protein-interacting conformation. Nature 455: 497–502. doi: 10.1038/nature07330
[14]	Hamm HE, Deretic D, Arendt A, et al. (1988) Site of G protein binding to rhodopsin mapped with synthetic peptides from the alpha subunit. Science 241: 832–835. doi: 10.1126/science.3136547
[15]	Pardon E, Laeremans T, Triest S, et al. (2014) A general protocol for the generation of Nanobodies for structural biology. Nat Protoc 9: 674–693. doi: 10.1038/nprot.2014.039
[16]	Carpenter B, Tate CG (2016) Engineering a minimal G protein to facilitate crystallisation of G protein-coupled receptors in their active conformation. Protein Eng Des Sel 29: 583–594.
[17]	Blankenship E, Vahedi-Faridi A, Lodowski DT (2015) The high-resolution structure of activated opsin reveals a conserved solvent network in the transmembrane region essential for activation. Structure 23: 2358–2364. doi: 10.1016/j.str.2015.09.015
[18]	Choe HW, Kim YJ, Park JH, et al. (2011) Crystal structure of metarhodopsin II. Nature 471: 651–655. doi: 10.1038/nature09789
[19]	Deupi X, Edwards P, Singhal A, et al. (2012) Stabilized G protein binding site in the structure of constitutively active metarhodopsin-II. Proc Natl Acad Sci USA 109: 119–124. doi: 10.1073/pnas.1114089108
[20]	Park JH, Morizumi T, Li Y, et al. (2013) Opsin, a structural model for olfactory receptors? Angew Chem Int Ed Engl 52: 11021–11024. doi: 10.1002/anie.201302374
[21]	Singhal A, Ostermaier MK, Vishnivetskiy SA, et al. (2013) Insights into congenital stationary night blindness based on the structure of G90D rhodopsin. EMBO Rep 14: 520–526. doi: 10.1038/embor.2013.44
[22]	Standfuss J, Edwards PC, D'Antona A, et al. (2011) The structural basis of agonist-induced activation in constitutively active rhodopsin. Nature 471: 656–660. doi: 10.1038/nature09795
[23]	Huang W, Manglik A, Venkatakrishnan AJ, et al. (2015) Structural insights into micro-opioid receptor activation. Nature 524: 315–321. doi: 10.1038/nature14886
[24]	Kruse AC, Ring AM, Manglik A, et al. (2013) Activation and allosteric modulation of a muscarinic acetylcholine receptor. Nature 504: 101–106. doi: 10.1038/nature12735
[25]	Rasmussen SG, Choi HJ, Fung JJ, et al. (2011) Structure of a nanobody-stabilized active state of the beta(2) adrenoceptor. Nature 469: 175–180. doi: 10.1038/nature09648
[26]	Ring AM, Manglik A, Kruse AC, et al. (2013) Adrenaline-activated structure of beta2-adrenoceptor stabilized by an engineered nanobody. Nature 502: 575–579. doi: 10.1038/nature12572
[27]	Weichert D, Kruse AC, Manglik A, et al. (2014) Covalent agonists for studying G protein-coupled receptor activation. Proc Natl Acad Sci USA 111: 10744–10748. doi: 10.1073/pnas.1410415111
[28]	Manglik A, Kobilka BK, Steyaert J (2017) Nanobodies to study G protein-coupled receptor structure and function. Annu Rev Pharmacol Toxicol 57: 19–37. doi: 10.1146/annurev-pharmtox-010716-104710
[29]	Nehme R, Carpenter B, Singhal A, et al. (2017) Mini-G proteins: Novel tools for studying GPCRs in their active conformation. PLoS One 12: e0175642. doi: 10.1371/journal.pone.0175642
[30]	Carpenter B, Nehme R, Warne T, et al. (2016) Structure of the adenosine A(2A) receptor bound to an engineered G protein. Nature 536: 104–107. doi: 10.1038/nature18966
[31]	Hanzal-Bayer M, Renault L, Roversi P, et al. (2002) The complex of Arl2-GTP and PDE delta: From structure to function. EMBO J 21: 2095–2106. doi: 10.1093/emboj/21.9.2095
[32]	Sunahara RK, Tesmer JJ, Gilman AG, et al. (1997) Crystal structure of the adenylyl cyclase activator Gsalpha. Science 278: 1943–1947. doi: 10.1126/science.278.5345.1943
[33]	Spiegel AM, Jr BP, Butrynski JE, et al. (1991) The G protein connection: Molecular basis of membrane association. Trends Biochem Sci 16: 338–341. doi: 10.1016/0968-0004(91)90139-M
[34]	Carpenter B, Tate CG (2017) Expression, purification and crystallisation of the adenosine A2A receptor bound to an engineered Mini G protein. Bio Protoc, 7.
[35]	Wan Q, Okashah N, Inoue A, et al. (2018) Mini G protein probes for active G protein-coupled receptors (GPCRs) in live cells. J Biol Chem 293: 7466–7473. doi: 10.1074/jbc.RA118.001975
[36]	Carpenter B, Tate CG (2017) Expression and purification of mini G proteins from escherichia coli. Bio Protoc 7: e2235.
[37]	Garcia-Nafria J, Lee Y, Bai X, et al. (2018) Cryo-EM structure of the adenosine A2A receptor coupled to an engineered heterotrimeric G protein. Elife 7: e35946. doi: 10.7554/eLife.35946
[38]	Garcia-Nafria J, Nehme R, Edwards PC, et al. (2018) Cryo-EM structure of the serotonin 5-HT1B receptor coupled to heterotrimeric Go. Nature 558: 620–623. doi: 10.1038/s41586-018-0241-9
[39]	Tsai CJ, Pamula F, Nehmé R, et al. (2018) Crystal structure of rhodopsin in complex with a mini-Go sheds light on the principles of G protein selectivity. Sci Adv 4: eaat7052. doi: 10.1126/sciadv.aat7052
[40]	Lebon G, Bennett K, Jazayeri A, et al. (2011) Thermostabilisation of an agonist-bound conformation of the human adenosine A(2A) receptor. J Mol Biol 409: 298–310. doi: 10.1016/j.jmb.2011.03.075
[41]	Lebon G, Warne T, Edwards PC, et al. (2011) Agonist-bound adenosine A2A receptor structures reveal common features of GPCR activation. Nature 474: 521–525. doi: 10.1038/nature10136
[42]	Warne T, Serrano-Vega MJ, Baker JG, et al. (2008) Structure of a beta1-adrenergic G-protein-coupled receptor. Nature 454: 486–491. doi: 10.1038/nature07101
[43]	Lebon G, Edwards PC, Leslie AG, et al. (2015) Molecular determinants of CGS21680 binding to the human adenosine A2A receptor. Mol Pharmacol 87: 907–915. doi: 10.1124/mol.114.097360
[44]	Lebon G, Warne T, Tate CG (2012) Agonist-bound structures of G protein-coupled receptors. Curr Opin Struct Biol 22: 482–490. doi: 10.1016/j.sbi.2012.03.007
[45]	Xu F, Wu H, Katritch V, et al. (2011) Structure of an agonist-bound human A2A adenosine receptor. Science 332: 322–327. doi: 10.1126/science.1202793
[46]	Ye L, Van EN, Zimmer M, et al. (2016) Activation of the A2A adenosine G-protein-coupled receptor by conformational selection. Nature 533: 265–268. doi: 10.1038/nature17668
[47]	Strege A, Carpenter B, Edwards PC, et al. (2017) Strategy for the thermostabilization of an agonist-bound GPCR coupled to a G protein. Method Enzymol 594: 243–264. doi: 10.1016/bs.mie.2017.05.014
[48]	Carpenter B, Lebon G (2017) Human adenosine A2A receptor: Molecular mechanism of ligand binding and activation. Front Pharmacol 8: 898. doi: 10.3389/fphar.2017.00898
[49]	Liang YL, Khoshouei M, Radjainia M, et al. (2017) Phase-plate cryo-EM structure of a class B GPCR-G-protein complex. Nature 546: 118–123. doi: 10.1038/nature22327
[50]	Liang YL, Khoshouei M, Deganutti G, et al. (2018) Cryo-EM structure of the active, Gs-protein complexed, human CGRP receptor. Nature 561: 492–497. doi: 10.1038/s41586-018-0535-y
[51]	Zhang Y, Sun B, Feng D, et al. (2017) Cryo-EM structure of the activated GLP-1 receptor in complex with a G protein. Nature 546: 248–253. doi: 10.1038/nature22394
[52]	Koehl A, Hu H, Maeda S, et al. (2018) Structure of the micro-opioid receptor-Gi protein complex. Nature 558: 547–552. doi: 10.1038/s41586-018-0219-7
[53]	Draper-Joyce CJ, Khoshouei M, Thal DM, et al. (2018) Structure of the adenosine-bound human adenosine A1 receptor-Gi complex. Nature 558: 559–563. doi: 10.1038/s41586-018-0236-6
[54]	Kang Y, Kuybeda O, de Waal PW, et al. (2018) Cryo-EM structure of human rhodopsin bound to an inhibitory G protein. Nature 558: 553–558. doi: 10.1038/s41586-018-0215-y
[55]	Khoshouei M, Radjainia M, Baumeister W, et al. (2017) Cryo-EM structure of haemoglobin at 3.2 A determined with the Volta phase plate. Nat Commun 8: 16099.
[56]	Renaud JP, Chari A, Ciferri C, et al. (2018) Cryo-EM in drug discovery: Achievements, limitations and prospects. Nat Rev Drug Discov 17: 471–492. doi: 10.1038/nrd.2018.77
[57]	Green SA, Holt BD, Liggett SB (1992) Beta 1- and beta 2-adrenergic receptors display subtype-selective coupling to Gs. Mol Pharmacol 41: 889–893.
[58]	Niesen FH, Berglund H, Vedadi M (2007) The use of differential scanning fluorimetry to detect ligand interactions that promote protein stability. Nat Protoc 2: 2212–2221. doi: 10.1038/nprot.2007.321
[59]	Yen HY, Hoi KK, Liko I, et al. (2018) PtdIns(4,5)P2 stabilizes active states of GPCRs and enhances selectivity of G-protein coupling. Nature 559: 423–427. doi: 10.1038/s41586-018-0325-6
[60]	Gales C, Rebois RV, Hogue M, et al. (2005) Real-time monitoring of receptor and G-protein interactions in living cells. Nat Methods 2: 177–184. doi: 10.1038/nmeth743
[61]	Hein P, Frank M, Hoffmann C, et al. (2005) Dynamics of receptor/G protein coupling in living cells. EMBO J 24: 4106–4114. doi: 10.1038/sj.emboj.7600870
[62]	Dixon AS, Schwinn MK, Hall MP, et al. (2016) NanoLuc complementation reporter optimized for accurate measurement of protein interactions in cells. ACS Chem Biol 11: 400–408. doi: 10.1021/acschembio.5b00753
[63]	Irannejad R, Pessino V, Mika D, et al. (2017) Functional selectivity of GPCR-directed drug action through location bias. Nat Chem Biol 13: 799–806. doi: 10.1038/nchembio.2389
[64]	Irannejad R, Tomshine JC, Tomshine JR, et al. (2013) Conformational biosensors reveal GPCR signalling from endosomes. Nature 495: 534–538. doi: 10.1038/nature12000
[65]	Kenakin T, Christopoulos A (2013) Signalling bias in new drug discovery: Detection, quantification and therapeutic impact. Nat Rev Drug Discov 12: 205–216.
[66]	Gentry PR, Sexton PM, Christopoulos A (2015) Novel allosteric modulators of G protein-coupled receptors. J Biol Chem 290: 19478–19488. doi: 10.1074/jbc.R115.662759
[67]	Skiba NP, Bae H, Hamm HE (1996) Mapping of effector binding sites of transducin alpha-subunit using G alpha t/G alpha i1 chimeras. J Biol Chem 271: 413–424. doi: 10.1074/jbc.271.1.413

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