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Crystal structure of an antiviral ankyrin targeting the HIV-1 capsid and molecular modeling of the ankyrin-capsid complex

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Abstract

Ankyrins are cellular repeat proteins, which can be genetically modified to randomize amino-acid residues located at defined positions in each repeat unit, and thus create a potential binding surface adaptable to macromolecular ligands. From a phage-display library of artificial ankyrins, we have isolated AnkGAG1D4, a trimodular ankyrin which binds to the HIV-1 capsid protein N-terminal domain (NTDCA) and has an antiviral effect at the late steps of the virus life cycle. In this study, the determinants of the AnkGAG1D4-NTDCA interaction were analyzed using peptide scanning in competition ELISA, capsid mutagenesis, ankyrin crystallography and molecular modeling. We determined the AnkGAG1D4 structure at 2.2 Å resolution, and used the crystal structure in molecular docking with a homology model of HIV-1 capsid. Our results indicated that NTDCA alpha-helices H1 and H7 could mediate the formation of the capsid-AnkGAG1D4 binary complex, but the interaction involving H7 was predicted to be more stable than with H1. Arginine-18 (R18) in H1, and R132 and R143 in H7 were found to be the key players of the AnkGAG1D4-NTDCA interaction. This was confirmed by R-to-A mutagenesis of NTDCA, and by sequence analysis of trimodular ankyrins negative for capsid binding. In AnkGAG1D4, major interactors common to H1 and H7 were found to be S45, Y56, R89, K122 and K123. Collectively, our ankyrin-capsid binding analysis implied a significant degree of flexibility within the NTDCA domain of the HIV-1 capsid protein, and provided some clues for the design of new antivirals targeting the capsid protein and viral assembly.

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References

  1. Cimarelli A, Darlix J-L (2002) Cell Mol Life Sci 59:1166

    Article  CAS  Google Scholar 

  2. Ganser BK, Li S, Klishko VY, Finch JT, Sundquist WI (1999) Science 283:80

    Article  CAS  Google Scholar 

  3. Ganser-Pornillos BK, Cheng A, Yeager M (2007) Cell 131:70

    Article  CAS  Google Scholar 

  4. Ganser-Pornillos BK, von Schwedler UK, Stray KM, Aiken C, Sundquist WI (2004) J Virol 78:2545

    Article  CAS  Google Scholar 

  5. Ganser-Pornillos BK, Yeager M, Sundquist WI (2008) Curr Opin Struct Biol 18:203

    Article  CAS  Google Scholar 

  6. von Schwedler UK, Stray KM, Garrus JE, Sundquist WI (2003) J Virol 77:5439

    Article  Google Scholar 

  7. Adamson CS, Freed EO (2007) Adv Pharmacol 55:347–387

    Article  CAS  Google Scholar 

  8. Freed EO (1998) Virology 251:1

    Article  CAS  Google Scholar 

  9. Freed EO (2002) J Virol 76:4679

    Article  CAS  Google Scholar 

  10. Göttlinger HG (2001) AIDS 15(Suppl. 5):S13

    Article  Google Scholar 

  11. Cardone G, Purdy JG, Cheng N, Craven RC, Steven AC (2009) Nature 457:694

    Article  CAS  Google Scholar 

  12. Darlix JL, Cristofari G, Rau M, Péchoux C, Berthoux L, Roques B (2000) Adv Pharmacol 48:345

    Article  CAS  Google Scholar 

  13. Fassati A (2012) Virus Res 170:15

    Article  CAS  Google Scholar 

  14. Pornillos O, Ganser-Pornillos BK, Kelly BN, Hua Y, Whitby FG, Stout CD, Sundquist WI, Hill CP, Yeager M (2009) Cell 137:1282

    Article  Google Scholar 

  15. Purdy JG, Flanagan JM, Ropson IJ, Craven RC (2009) J Mol Biol 389:438

    Article  CAS  Google Scholar 

  16. Amstutz P, Binz HK, Parizek P, Stumpp MT, Kohl A, Grütter MG, Forrer P, Plückthun A (2005) J Biol Chem 280:24715

    Article  CAS  Google Scholar 

  17. Binz HK, Stumpp MT, Forrer P, Amstutz P, Pluckthun A (2003) J Mol Biol 332:489

    Article  CAS  Google Scholar 

  18. Kummer L, Parizek P, Rube P, Millgramm B, Prinz A, Mittl PR, Kaufholz M, Zimmermann B, Herberg FW, Plückthun A (2012) Proc Natl Acad Sci USA 109:E2248

    Article  CAS  Google Scholar 

  19. Li J, Mahajan A, Tsai MD (2006) Biochemistry 45:15168

    Article  CAS  Google Scholar 

  20. Mosavi LK, Cammett TJ, Desrosiers DC, Peng ZY (2004) Protein Sci 13:1435

    Article  CAS  Google Scholar 

  21. Schweizer A, Roschitzki-Voser H, Amstutz P, Briand C, Gulotti-Georgieva M, Prenosil E, Binz HK, Capitani G, Baici A, Pluckthun A, Grutter MG (2007) Structure 15:625

    Article  CAS  Google Scholar 

  22. Schweizer A, Rusert P, Berlinger L, Ruprecht CR, Mann A, Corthésy S, Turville SG, Aravantinou M, Fischer M, Robbiani M, Amstutz P, Trkola A (2008) PLoS Pathog 4:e1000109

    Article  Google Scholar 

  23. Sedgwick SG, Smerdon SJ (1999) Trends Biochem Sci 24:311

    Article  CAS  Google Scholar 

  24. Zahnd C, Wyler E, Schwenk JM, Steiner D, Lawrence MC, McKern NM, Pecorari F, Ward CW, Joos TO, Plückthun A (2007) J Mol Biol 369:1015

    Article  CAS  Google Scholar 

  25. Tamaskovic R, Simon M, Stefan N, Schwill M, Pluckthun A (2012) Methods Enzymol 503:101–134

    Article  CAS  Google Scholar 

  26. Nangola S, Urvoas A, Valerio-Lepiniec M, Khamaikawin W, Sakkhachornphop S, Hong SS, Boulanger P, Minard P, Tayapiwatana C (2012) Retrovirology 9:17

    Article  CAS  Google Scholar 

  27. Buranapraditkun S, Hempel U, Pitakpolrat P, Allgaier RL, Thantivorasit P, Lorenzen SI, Sirivichayakul S, Hildebrand WH, Altfeld M, Brander C, Walker BD, Phanuphak P, Hansasuta P, Rowland-Jones SL, Allen TM, Ruxrungtham K (2011) PLoS ONE 6:e23603

    Article  CAS  Google Scholar 

  28. DaFonseca S, Blommaert A, Coric P, Hong SS, Bouaziz S, Boulanger P (2007) Antiviral Ther 12:1185

    CAS  Google Scholar 

  29. Gay B, Tournier J, Chazal N, Carrière C, Boulanger P (1998) Virology 247:160

    Article  CAS  Google Scholar 

  30. DaFonseca S, Coric P, Gay B, Hong SS, Bouaziz S, Boulanger P (2008) Virol J 5:162

    Article  Google Scholar 

  31. Kitidee K, Nangola S, Gonzalez G, Boulanger P, Tayapiwatana C, Hong SS (2010) BMC Biotechnol 10:80

    Article  Google Scholar 

  32. Otwinowski Z, Minor W (1997) Methods Enzymol 276:307

    Article  CAS  Google Scholar 

  33. Monroe N, Sennhauser G, Seeger MA, Briand C, Grütter MG (2011) J Struct Biol 174:269

    Article  CAS  Google Scholar 

  34. Winn MD, Ballard CC, Cowtan KD, Dodson EJ, Emsley P, Evans PR, Keegan RM, Krissinel EB, Leslie AG, Mccoy A, McNicholas SJ, Murshudov GN, Pannu NS, Potterton EA, Powell HR, Read RJ, Vagin A, Wilson KS (2011) Acta Cryst D 67:235

    Article  CAS  Google Scholar 

  35. Emsley P, Cowtan KD (2004) Acta Crystallogr D Biol Crystallogr 60:2126

    Article  Google Scholar 

  36. Laskowski RA, MacArthur MW, Moss DS, Thornton JM (1993) J Appl Crystallogr 26:283

    Article  CAS  Google Scholar 

  37. Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ (1997) Nucleic Acids Res 25:3389

    Article  CAS  Google Scholar 

  38. Shin R, Tzou YM, Krishna NR (2011) Biochemistry 50:9457

    Article  CAS  Google Scholar 

  39. Schwede T, Kopp J, Guex N, Peitsch MC (2003) Nucl Acids Res 31:3381

    Article  CAS  Google Scholar 

  40. Chen R, Li L, Weng Z (2003) Proteins 52:80

    Article  CAS  Google Scholar 

  41. Chen R, Weng Z (2003) Proteins 51:397

    Article  CAS  Google Scholar 

  42. Hwang H, Vreven T, Pierce BG, Hung JH, Weng Z (2010) Proteins 78:3104

    Article  CAS  Google Scholar 

  43. Li L, Chen R, Weng Z (2003) Proteins 53:693

    Article  CAS  Google Scholar 

  44. Wisitponchai T, Nimmanpipug P, Lee VS, Tayapiwatana C (2013) Chiang Mai J Sci 40:232

    Google Scholar 

  45. Cressey R, Pimpa S, Chewaskulyong B, Lertprasertsuke N, Saeteng S, Tayapiwatana C, Kasinrerk W (2008) BMC Biotechnol 8:16

    Article  Google Scholar 

  46. Gitti RK, Lee BM, Walker J, Summers MF, Yoo S, Sundquist WI (1996) Science 273:231

    Article  CAS  Google Scholar 

  47. Apolloni A, Lin M-H, Sivakumaran H, Li D, Kershaw MHR, Harrich D (2013) Hum Gene Ther 24:270

    Article  CAS  Google Scholar 

  48. Checkley MA, Luttge BG, Soheilian F, Nagashima K, Freed EO (2010) Virology 400:137

    Article  CAS  Google Scholar 

  49. Fackler OT, D’Aloja P, Baur AS, Federico M, Peterlin BM (2001) J Virol 75:6601

    Article  CAS  Google Scholar 

  50. Trono D, Feinberg MB, Baltimore D (1989) Cell 59:113

    Article  CAS  Google Scholar 

  51. Walker RC Jr, Khan MA, Kao S, Goila-Gaur R, Miyagi E, Strebel K (2010) J Virol 84:5201

    Article  CAS  Google Scholar 

  52. Neagu MR, Ziegler P, Pertel T, Strambio-De-Castillia C, Grütter C, Martinetti G, Mazzucchelli L, Grütter MG, Manz MG, Luban J (2009) J Clin Invest 119:3035

    Article  CAS  Google Scholar 

  53. Tang C, Loeliger E, Kinde I, Kyere S, Mayo K, Barklis E, Sun Y, Huang M, Summers MF (2003) J Mol Biol 327:1013

    Article  CAS  Google Scholar 

  54. Bartonova V, Igonet S, Sticht J, Glass B, Habermann A, Vaney MC, Sehr P, Lewis J, Rey FA, Kräusslich HG (2008) J Biol Chem 283:32024

    Article  CAS  Google Scholar 

  55. Sticht J, Humbert M, Findlow S, Bodem J, Muller B, Dietrich U, Werner J, Kräusslich H-G (2005) Nat Struct Mol Biol 12:671

    Article  CAS  Google Scholar 

  56. Blair WS, Pickford C, Irving SL, Brown DG, Anderson M, Bazin R, Cao J, Ciaramella G, Isaacson J, Jackson L, Hunt R, Kjerrstrom A, Nieman JA, Patick AK, Perros M, Scott AD, Whitby K, Wu H, Butler SL (2010) PLoS Pathog 6:e1001220

    Article  Google Scholar 

  57. Shi J, Zhou J, Shah VB, Aiken C, Whitby K (2011) J Virol 85:542

    Article  CAS  Google Scholar 

  58. Nangola S, Minard P, Tayapiwatana C (2010) Protein Expr Purif 74:156

    Article  CAS  Google Scholar 

  59. Saint-Remy JM (1997) Toxicology 119:77

    Article  CAS  Google Scholar 

  60. Lenstra JA, Kusters JG, van der Zeijst BA (1990) Arch Virol 110:1

    Article  CAS  Google Scholar 

  61. Stanley KK, Herz J (1987) EMBO J 6:1951–1957

    CAS  Google Scholar 

  62. van Regenmortel MH (1989) Immunol Today 10:266

    Article  Google Scholar 

  63. Sennhauser G, Grütter MG (2008) Structure 16:1443

    Article  CAS  Google Scholar 

  64. Zhao Y, Chen Y, Schutkowski M, Fischer G, Ke H (1997) Structure 5:139

    Article  CAS  Google Scholar 

  65. Price AJ, Fletcher AJ, Schaller T, Elliott T, Lee K, KewalRamani VN, Chin JW, Towers GJ, James LC (2012) PLoS Pathog 8:e1002896

    Article  CAS  Google Scholar 

  66. Li G, Verheyen J, Rhee S-Y, Voet A, Vandamme A-M, Theys K (2013) Retrovirology 10:126

    Article  Google Scholar 

  67. Goudreau N, Coulombe R, Faucher AM, Grand-Maitre C, Lacoste JE, Lemke CT, Malenfant E, Bousquet Y, Fader L, Simoneau B, Mercier JF, Titolo S, Mason SW (2013) Chem Med Chem 8:405

    Article  CAS  Google Scholar 

  68. Lamorte L, Titolo S, Lemke CT, Goudreau N, Mercier JF, Wardrop E, Shah VB, von Schwedler UK, Langelier C, Banik SS, Aiken C, Sundquist WI, Mason SW (2013) Antimicrob Agents Chemother 57:4622

    Article  CAS  Google Scholar 

  69. Lemke CT, Titolo S, von Schwedler UK, Goudreau N, Mercier JF, Wardrop E, Faucher AM, Coulombe R, Banik SS, Fader L, Gagnon A, Kawai SH, Rancourt J, Tremblay M, Yoakim C, Simoneau B, Archambault J, Sundquist WI, Mason SW (2012) J Virol 86:6643

    Article  CAS  Google Scholar 

  70. Kelly BN, Kyere S, Kinde I, Tang C, Howard BR, Robinson H, Sundquist WI, Summers MF, Hill C (2007) J Mol Biol 373:355

    Article  CAS  Google Scholar 

  71. Rein A, Datta SA, Jones CP, Musier-Forsyth K (2011) Trends Biochem Sci 36:373

    CAS  Google Scholar 

  72. Datta SA, Heinrich F, Raghunandan S, Krueger S, Curtis JE, Rein A, Nanda H (2010) J Mol Biol 406:205

    Article  Google Scholar 

  73. Engelman A, Cherepanov P (2012) Nat Rev Microbiol 10:279

    Article  CAS  Google Scholar 

  74. Li L, Oliveira NMM, Cheney KM, Pade C, Dreja H, Bergin A-MH, Borgdorff V, Beach DH, Bishop CL, Dittmar MT, McKnight A (2011) Retrovirology 8:94

    Article  Google Scholar 

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Acknowledgments

The work in Thailand was supported by the National Research University project under the Thailand’s Office of the Commission on Higher Education, the Research Chair Grant of National Sciences and the Technology Development Agency at the Faculty of Associated Medical Sciences, Chiang Mai University, and the Thailand Research Fund. The work in Lyon was supported by the French National Agency for Research on AIDS and Viral Hepatitis (Inserm-ANRS contract No. 11344/2011-2013). We are grateful to thank Prof. Dr. Kiat Ruxrungtham for his kind gift of synthetic pentadecapeptides, and the NIH AIDS Research and Reference Reagent Program for providing us with the pNL4-3 plasmid. The authors would like to thank the National Electronics and Computer Technology (NECTEC) for Discovery Studio software and also thank the University of Malaya for UMRG (Grant No. RP002-2012D) and FRGS (FP008-2012A), and for its support with computational work. We are also indebted to the supporting staffs at beamline BL13B1 at NSRRC and beamline BL12B2 at SPring-8 for the technical assistance. The crystallographic work was supported in part by National Science Council Grants 101-2628-B-213-001-MY4 and NSRRC Grant 1013RSB02 to C.-J. Chen.

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Authors and Affiliations

  1. Division of Clinical Immunology, Department of Medical Technology, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai, 50200, Thailand

    Warachai Praditwongwan, Somphot Saoin & Chatchai Tayapiwatana

  2. Biomedical Technology Research Unit, National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency at the Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai, 50200, Thailand

    Warachai Praditwongwan, Somphot Saoin & Chatchai Tayapiwatana

  3. Life Science Group, Scientific Research Division, National Synchrotron Radiation Research Center, 101 Hsin-Ann Road, Hsinchu, 30076, Taiwan

    Phimonphan Chuankhayan & Chun-Jung Chen

  4. Department of Physics, National Tsing Hua University, Hsinchu, 30013, Taiwan

    Chun-Jung Chen

  5. Institute of Biotechnology, National Cheng Kung University, Tainan, 701, Taiwan

    Chun-Jung Chen

  6. Biomedical Engineering Center, Faculty of Engineering, Chiang Mai University, Chiang Mai, 50200, Thailand

    Tanchanok Wisitponchai

  7. Computational Simulation Modelling Laboratory (CSML), Department of Chemistry and Center of Excellence for Innovation in Chemistry and Materials Science Research Center, Faculty of Science, Chiang Mai University, Chiang Mai, 50200, Thailand

    Vannajan Sanghiran Lee

  8. Department of Chemistry, Faculty of Science, University of Malaya, 50603, Kuala Lumpur, Malaysia

    Vannajan Sanghiran Lee

  9. Division of Clinical Immunology and Transfusion Sciences, Department of Medical Technology, School of Allied Health Sciences, University of Phayao, Phayao, 56000, Thailand

    Sawitree Nangola

  10. Laboratory of Retrovirus and Comparative Pathology, UMR-754, University Lyon 1, 50, Avenue Tony Garnier, 69366, Lyon Cedex 07, France

    Saw See Hong & Pierre Boulanger

  11. UMR-754, Retrovirus and Comparative Pathology, INRA, 50, Avenue Tony Garnier, 69366, Lyon Cedex 07, France

    Saw See Hong & Pierre Boulanger

  12. Institut National de la Santé et de la Recherche Médicale (INSERM), 101, Rue de Tolbiac, 75013, Paris, France

    Saw See Hong

  13. Institut de Biochimie et de Biophysique Moléculaire et Cellulaire (IBBMC) UMR-8619, Université de Paris-Sud et CNRS, 91405, Orsay Cedex, France

    Philippe Minard

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  1. Warachai Praditwongwan
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  2. Phimonphan Chuankhayan
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  5. Vannajan Sanghiran Lee
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  6. Sawitree Nangola
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  7. Saw See Hong
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  8. Philippe Minard
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  9. Pierre Boulanger
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  10. Chun-Jung Chen
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Corresponding authors

Correspondence to Pierre Boulanger, Chun-Jung Chen or Chatchai Tayapiwatana.

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Warachai Praditwongwan and Phimonphan Chuankhayan have contributed equally.

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Praditwongwan, W., Chuankhayan, P., Saoin, S. et al. Crystal structure of an antiviral ankyrin targeting the HIV-1 capsid and molecular modeling of the ankyrin-capsid complex. J Comput Aided Mol Des 28, 869–884 (2014). https://doi.org/10.1007/s10822-014-9772-9

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