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Molecular modeling of Trypanosoma cruzi glutamate cysteine ligase and investigation of its interactions with glutathione

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Abstract

Trypanosoma cruzi glutamate cysteine ligase (TcGCL) is considered a potential drug target to develop novel antichagasic drugs. We have used a variety of computational methods to investigate the interactions between TcGCL with Glutathione (GSH). The three-dimensional structure of TcGCL was constructed by comparative modeling methods using the Saccharomyces cerevisiae glutamate cysteine ligase as template. Molecular dynamics simulations were used to validate the TcGCL model and to analyze the molecular interactions with GSH. Using RMSD clustering, the most prevalent GSH binding modes were identified paying attention to the residues involved in the molecular interactions. The GSH binding modes were used to propose pharmacophore models that can be exploited in further studies to identify novel antichagasic compounds.

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References

  1. Moncayo A, Silveira AC (2009) Current epidemiological trends for Chagas disease in Latin America and future challenges in epidemiology, surveillance and health policy. Mem Inst Oswaldo Cruz 104(Suppl 1):17–30

    Google Scholar 

  2. Abbott JJ, Pei J, Ford JL, Qi Y, Grishin VN, Pitcher LA, Phillips MA, Grishin NV (2001) Structure prediction and active site analysis of the metal binding determinants in gamma -glutamylcysteine synthetase. J Biol Chem 276:42099–42107

    Article  CAS  Google Scholar 

  3. Brekken DL, Phillips MA (1998) Trypanosoma brucei gamma-glutamylcysteine synthetase. Characterization of the kinetic mechanism and the role of Cys-319 in cystamine inactivation. J Biol Chem 273:26317–26322

    Article  CAS  Google Scholar 

  4. Huynh TT, Huynh VT, Harmon MA, Phillips MA (2003) Gene Knockdown of γ-Glutamylcysteine Synthetase by RNAi in the Parasitic Protozoa Trypanosoma brucei Demonstrates That It Is an Essential Enzyme. J Biol Chem 278:39794–39800

    Article  CAS  Google Scholar 

  5. Krzywanski DM, Dickinson DA, Iles KE, Wigley AF, Franklin CC, Liu RM, Kavanagh TJ, Forman HJ (2004) Variable regulation of glutamate cysteine ligase subunit proteins affects glutathione biosynthesis in response to oxidative stress. Arch Biochem Biophys 423:116–125

    Article  CAS  Google Scholar 

  6. Hiratake J (2005) Enzyme inhibitors as chemical tools to study enzyme catalysis: rational design, synthesis, and applications. Chem Rec 5:209–228

    Article  CAS  Google Scholar 

  7. Ariyanayagam MR, Oza SL, Mehlert A, Fairlamb AH (2003) Bis(glutathionyl)spermine and other novel trypanothione analogues in Trypanosoma cruzi. J Biol Chem 278:27612–27619

    Article  CAS  Google Scholar 

  8. Oza SL, Tetaud E, Ariyanayagam MR, Warnon SS, Fairlamb AH (2002) A single enzyme catalyses formation of Trypanothione from glutathione and spermidine in Trypanosoma cruzi. J Biol Chem 277:35853–35861

    Article  CAS  Google Scholar 

  9. Wyllie S, Oza SL, Patterson S, Spinks D, Thompson S, Fairlamb AH (2009) Dissecting the essentiality of the bifunctional trypanothione synthetase-amidase in Trypanosoma brucei using chemical and genetic methods. Mol Microbiol 74:529–540

    Article  CAS  Google Scholar 

  10. Copley SD, Dhillon JK (2002) Lateral gene transfer and parallel evolution in the history of glutathione biosynthesis genes. Genome Biol 3:research0025

    Google Scholar 

  11. Biterova EI, Barycki JJ (2010) Structural basis for feedback and pharmacological inhibition of Saccharomyces cerevisiae glutamate cysteine ligase. J Biol Chem 285:14459–14466

    Article  CAS  Google Scholar 

  12. May MJ, Leaver CJ (1994) Arabidopsis thaliana gamma-glutamylcysteine synthetase is structurally unrelated to mammalian, yeast, and Escherichia coli homologs. Proc Natl Acad Sci USA 91:10059–10063

    Article  CAS  Google Scholar 

  13. Hothorn M, Wachter A, Gromes R, Stuwe T, Rausch T, Scheffzek K (2006) Structural basis for the redox control of plant glutamate cysteine ligase. J Biol Chem 281:27557–27565

    Article  CAS  Google Scholar 

  14. Hibi T, Nii H, Nakatsu T, Kimura A, Kato H, Hiratake J, Oda J (2004) Crystal structure of gamma-glutamylcysteine synthetase: insights into the mechanism of catalysis by a key enzyme for glutathione homeostasis. Proc Natl Acad Sci USA 101:15052–15057

    Article  CAS  Google Scholar 

  15. Hamilton D, Wu JH, Batist G (2007) Structure-based identification of novel human gamma-glutamylcysteine synthetase inhibitors. Mol Pharmacol 71:1140–1147

    Article  CAS  Google Scholar 

  16. Griffith OW (1982) Mechanism of action, metabolism, and toxicity of buthionine sulfoximine and its higher homologs, potent inhibitors of glutathione synthesis. J Biol Chem 257:13704–13712

    CAS  Google Scholar 

  17. Griffith OW, Meister A (1979) Potent and specific inhibition of glutathione synthesis by buthionine sulfoximine (S-n-butyl homocysteine sulfoximine). J Biol Chem 254:7558–7560

    CAS  Google Scholar 

  18. Griffith OW, Anderson ME, Meister A (1979) Inhibition of glutathione biosynthesis by prothionine sulfoximine (S-n-propyl homocysteine sulfoximine), a selective inhibitor of gamma-glutamylcysteine synthetase. J Biol Chem 254:1205–1210

    CAS  Google Scholar 

  19. Faundez M, Pino L, Letelier P, Ortiz C, Lopez R, Seguel C, Ferreira J, Pavani M, Morello A, Maya JD (2005) Buthionine sulfoximine increases the toxicity of nifurtimox and benznidazole to Trypanosoma cruzi. Antimicrob Agents Chemother 49:126–130

    Article  CAS  Google Scholar 

  20. Moncada C, Repetto Y, Aldunate J, Letelier ME, Morello A (1989) Role of glutathione in the susceptibility of Trypanosoma cruzi to drugs. Comp Biochem Physiol C 94:87–91

    Article  CAS  Google Scholar 

  21. Repetto Y, Opazo E, Maya JD, Agosin M, Morello A (1996) Glutathione and trypanothione in several strains of Trypanosoma cruzi: effect of drugs. Comp Biochem Physiol B Biochem Mol Biol 115:281–285

    Article  CAS  Google Scholar 

  22. Maya JD, Cassels BK, Iturriaga-Vasquez P, Ferreira J, Faundez M, Galanti N, Ferreira A, Morello A (2007) Mode of action of natural and synthetic drugs against Trypanosoma cruzi and their interaction with the mammalian host. Comp Biochem Physiol A Mol Integr Physiol 146:601–620

    Article  Google Scholar 

  23. Harvison PJ, Kalman TI (1992) Synthesis and biological activity of novel folic acid analogues: pteroyl-S-alkylhomocysteine sulfoximines. J Med Chem 35:1227–1233

    Article  CAS  Google Scholar 

  24. Abbott JJ, Ford JL, Phillips MA (2002) Substrate binding determinants of Trypanosoma brucei gamma-glutamylcysteine synthetase. Biochemistry 41:2741–2750

    Article  CAS  Google Scholar 

  25. Janowiak BE, Griffith OW (2005) Glutathione synthesis in Streptococcus agalactiae. One protein accounts for gamma-glutamylcysteine synthetase and glutathione synthetase activities. J Biol Chem 280:11829–11839

    Article  CAS  Google Scholar 

  26. Jez JM, Cahoon RE, Chen S (2004) Arabidopsis thaliana glutamate-cysteine ligase: functional properties, kinetic mechanism, and regulation of activity. J Biol Chem 279:33463–33470

    Article  CAS  Google Scholar 

  27. Huynh TT, Huynh VT, Harmon MA, Phillips MA (2003) Gene knockdown of gamma-glutamylcysteine synthetase by RNAi in the parasitic protozoa Trypanosoma brucei demonstrates that it is an essential enzyme. J Biol Chem 278:39794–39800

    Article  CAS  Google Scholar 

  28. Ashida H, Sawa Y, Shibata H (2005) Cloning, biochemical and phylogenetic characterizations of gamma-glutamylcysteine synthetase from Anabaena sp. PCC 7120. Plant Cell Physiol 46:557–562

    Article  CAS  Google Scholar 

  29. Drew R, Miners JO (1984) The effects of buthionine sulphoximine (BSO) on glutathione depletion and xenobiotic biotransformation. Biochem Pharmacol 33:2989–2994

    Article  CAS  Google Scholar 

  30. Sali A, Blundell TL (1993) Comparative protein modelling by satisfaction of spatial restraints. J Mol Biol 234:779–815

    Article  CAS  Google Scholar 

  31. The UniProt Consortium (2009) The Universal Protein Resource (UniProt) 2009. Nucl Acids Res 37:D169–D174

    Article  Google Scholar 

  32. Green JR, Korenberg MJ, Aboul-Magd MO (2009) PCI-SS: MISO dynamic nonlinear protein secondary structure prediction. BMC Bioinforma 10:222

    Article  Google Scholar 

  33. Brooks BR, Bruccoleri RE, Olafson BD, States DJ, Swaminathan S, Karplus M (1983) CHARMM: A program for macromolecular energy, minimization, and dynamics calculations. J Comput Chem 4:187–217

    Article  CAS  Google Scholar 

  34. Lovell SC, Davis IW, Arendall WB 3rd, de Bakker PI, Word JM, Prisant MG, Richardson JS, Richardson DC (2003) Structure validation by Calpha geometry: phi, psi and Cbeta deviation. Proteins 50:437–450

    Article  CAS  Google Scholar 

  35. Luthy R, Bowie JU, Eisenberg D (1992) Assessment of protein models with three-dimensional profiles. Nature 356:83–85

    Article  CAS  Google Scholar 

  36. Wiederstein M, Sippl MJ (2007) ProSA-web: interactive web service for the recognition of errors in three-dimensional structures of proteins. Nucleic Acids Res 35:W407–W410

    Article  Google Scholar 

  37. Wallner B, Elofsson A (2003) Can correct protein models be identified? Protein Sci 12:1073–1086

    Article  CAS  Google Scholar 

  38. Honig B, Nicholls A (1995) Classical electrostatics in biology and chemistry. Science 268:1144–1149

    Article  CAS  Google Scholar 

  39. Humphrey W, Dalke A, Schulten K (1996) VMD: Visual molecular dynamics. J Mol Graph 14:33–38

    Article  CAS  Google Scholar 

  40. Phillips JC, Braun R, Wang W, Gumbart J, Tajkhorshid E, Villa E, Chipot C, Skeel RD, Kale L, Schulten K (2005) Scalable molecular dynamics with NAMD. J Comput Chem 26:1781–1802

    Article  CAS  Google Scholar 

  41. Mackerell AD Jr (2004) Empirical force fields for biological macromolecules: overview and issues. J Comput Chem 25:1584–1604

    Article  CAS  Google Scholar 

  42. MacKerell AD, Bashford D, Bellott DRL, Evanseck JD, Field MJ, Fischer S, Gao J, Guo H, Ha S, Joseph-McCarthy D, Kuchnir L, Kuczera K, Lau FTK, Mattos C, Michnick S, Ngo T, Nguyen DT, Prodhom B, Reiher WE, Roux B, Schlenkrich M, Smith JC, Stote R, Straub J, Watanabe M, Wiorkiewicz-Kuczera J, Yin D, Karplus M (1998) All-Atom Empirical Potential for Molecular Modeling and Dynamics Studies of Proteins. J Phys Chem B 102:3586–3616

    Article  CAS  Google Scholar 

  43. Jorgensen WL, Chandrasekhar J, Madura JD, Impey RW, Klein ML (1983) Comparison of simple potential functions for simulating liquid water. J Chem Phys 79:926–935

    Article  CAS  Google Scholar 

  44. Darden T, York D, Pedersen L (1993) Particle mesh Ewald: An N [center-dot] log(N) method for Ewald sums in large systems. J Chem Phys 98:10089–10092

    Article  CAS  Google Scholar 

  45. Essmann U, Perera L, Berkowitz ML, Darden T, Lee H, Pedersen LG (1995) A smooth particle mesh Ewald method. J Chem Phys 103:8577–8593

    Article  CAS  Google Scholar 

  46. Feller SE, Zhang Y, Pastor RW, Brooks BR (1995) Constant pressure molecular dynamics simulation: The Langevin piston method. J Chem Phys 103:4613–4621

    Article  CAS  Google Scholar 

  47. Ryckaert J-P, Ciccotti G, Berendsen HJC (1977) Numerical integration of the cartesian equations of motion of a system with constraints: molecular dynamics of n-alkanes. J Chem Phys 23:327–341

    CAS  Google Scholar 

  48. Seeber M, Cecchini M, Rao F, Settanni G, Caflisch A (2007) Wordom: a program for efficient analysis of molecular dynamics simulations. Bioinformatics 23:2625–2627

    Article  CAS  Google Scholar 

  49. Berman HM, Westbrook J, Feng Z, Gilliland G, Bhat TN, Weissig H, Shindyalov IN, Bourne PE (2000) The Protein Data Bank. Nucleic Acids Res 28:235–242

    Article  CAS  Google Scholar 

  50. Frishman D, Argos P (1995) Knowledge-based protein secondary structure assignment. Proteins 23:566–579

    Article  CAS  Google Scholar 

  51. Ferre F, Clote P (2005) DiANNA: a web server for disulfide connectivity prediction. Nucleic Acids Res 33:W230–W232

    Article  CAS  Google Scholar 

  52. Lueder DV, Phillips MA (1996) Characterization of Trypanosoma brucei γ-Glutamylcysteine Synthetase, an Essential Enzyme in the Biosynthesis of Trypanothione (Diglutathionylspermidine). J Biol Chem 271:17485–17490

    Article  CAS  Google Scholar 

  53. Brekken DL, Phillips MA (1998) Trypanosoma brucei Gamma-Glutamylcysteine Synthetase. J Biol Chem 273:26317–26322

    Article  CAS  Google Scholar 

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Acknowledgments

The authors would like to thank Fondo Nacional de Desarrollo Científico y Tecnológico (FONDECYT, Chile) project Nº 11085027 to COS for financial support. RAT acknowledges Programa Bicentenario de Ciencia y Tecnología del Proyecto de Inserción (PSD70). TPA acknowledge to Proyectos de Financiamiento Basal PFB03 and PFB16, and to Millenium Institute Centro Interdisciplinario de Neurociencias de Valparaíso (CINV). Molecular dynamics simulations were performed at the National Laboratory for High Performance Computing (NLHPC ECM-02) supercomputing infrastructure: Powered@NLHPC. CFL, RAS and PT are PhD fellows from Comisión Nacional de Investigación Científica y Tecnológica (CONICYT, Chile).

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Authors and Affiliations

  1. Departamento de Farmacia, Facultad de Quimica, P. Universidad Catolica de Chile, Av Vicuña Mackenna 4860, Macul-Santiago, Chile

    Carlos F. Lagos

  2. Departamento de Ciencias Biologicas, Facultad de Ciencias Biologicas, Universidad Andres Bello, Av Republica 217, Santiago, Chile

    Carlos F. Lagos & Raul Araya-Secchi

  3. Computational Biology Lab (DLab), Centro de Modelamiento Matematico (CMM), Facultad de Ciencias Fisicas y Matematicas, Universidad de Chile, Santiago, Chile

    Raul Araya-Secchi & Tomás Pérez-Acle

  4. Fundacion Ciencia para la Vida, Zañartu 1482, Ñuñoa, Santiago, Chile

    Tomás Pérez-Acle

  5. Centro Interdisciplinario de Neurociencias de Valparaiso (CINV), Pasaje Harrington 287, Playa Ancha, Valparaiso, Chile

    Tomás Pérez-Acle

  6. Departamento de Quimica Organica, Facultad de Quimica, P. Universidad Catolica de Chile, Av Vicuña Mackenna 4860, Macul- Santiago, Chile

    Pablo Thomas, Ricardo A. Tapia & Cristian O. Salas

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  1. Carlos F. Lagos
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  2. Raul Araya-Secchi
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  3. Pablo Thomas
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  4. Tomás Pérez-Acle
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  5. Ricardo A. Tapia
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  6. Cristian O. Salas
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Corresponding author

Correspondence to Cristian O. Salas.

Additional information

Carlos F. Lagos and Raul Araya-Secchi have contributed equally to this work.

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Supplementary Fig. 1

Sequence alignment between Trypanosoma cruzi (Tc) and Trypanosoma brucei (Tb) GCLs. Sequences were retrieved from Uniprot Database (http://www.uniprot.org/). Sequence identity is 67.7% and sequence similarity is 81.6%. Red asterisks denote residues reported in site directed mutagenesis referred in the text. (DOC 21946 kb)

Supplementary Fig. 2

A) Procheck generated Ramachandran plots and statistics for template and modeled TcGCL protein. B) Verify-3D profiles obtained using the SAVES server. (DOC 19498 kb)

Supplementary Fig. 3

Prosa-web results for template protein (PDB 3LVW in upper panel) and modeled TcGCL (lower panel). A) Z-plot graphs and B) e-plot graph. (DOC 6921 kb)

Supplementary 4

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Lagos, C.F., Araya-Secchi, R., Thomas, P. et al. Molecular modeling of Trypanosoma cruzi glutamate cysteine ligase and investigation of its interactions with glutathione. J Mol Model 18, 2055–2064 (2012). https://doi.org/10.1007/s00894-011-1224-z

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  • DOI: https://doi.org/10.1007/s00894-011-1224-z

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