Transit of Procaspase-9 towards its activation. New mechanistic insights from molecular dynamics simulations

  • Humberto Gasperin-Sánchez
  • Claudia G. Benítez-Cardoza
  • Luis A. Caro-Gómez
  • Jorge L. Rosas-Trigueros
  • Absalom Zamorano-CarrilloEmail author
Original Paper


Caspases are cysteine proteases that perform a wide variety of roles in lethal intracellular signaling and cell-death regulation. Caspase-9, the primary initiator caspase of the intrinsic apoptotic pathway, is produced as a scarcely active zymogen (Procaspase-9). Here, we describe, for the first time, at the atomistic level, conformational changes which might be correlated to the activation of Procaspase-9. Molecular dynamics simulations performed at two temperatures (310 and 410 K) provide insights about the conformational space and the time-course evolution of the geometrical and structural characteristics of Procaspase-9. At both temperatures studied, the extremal globular domains of the protein approach each other, contracting the disordered region. In both temperatures, the compact conformations hide more than 40 nm2 (about 20% of the total solvent-accessible surface area), and their radius of gyration are reduced by about 40% from the original values. At each temperature, the pathway of contraction is different, as well as the compact structures reached. In consequence, the network of stabilizing interactions at the final conformations is dissimilar. Both final conformations were evaluated in their structural compatibility with the activation models described so far. In this work, we describe mechanistically how and why the activation of Procaspase-9 is favored by apoptosome recruitment via the Caspase Activation Recruitment Domain (CARD), as it has been proposed recently by in vitro experiments.


Molecular dynamics Procaspase-9 Caspase Activation Apoptosome 


Supplementary material

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  1. 1.
    Shalini S, Dorstyn L, Dawar S, Kumar S (2015) Old, new and emerging functions of caspases. Cell Death Differ 22(4):526–539. CrossRefPubMedGoogle Scholar
  2. 2.
    Shi Y (2004) Caspase activation, inhibition, and reactivation: a mechanistic view. Protein Sci 13(8):1979–1987. CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Wu CC, Lee S, Malladi S, Chen MD, Mastrandrea NJ, Zhang Z, Bratton SB (2016) The Apaf-1 apoptosome induces formation of caspase-9 homo- and heterodimers with distinct activities. Nat Commun 7:13565. CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Wu CC, Bratton SB (2017) Caspase-9 swings both ways in the apoptosome. Mol Cell Oncol 4(2):e1281865. CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Wurstle ML, Rehm M (2014) A systems biology analysis of apoptosome formation and apoptosis execution supports allosteric procaspase-9 activation. J Biol Chem 289(38):26277–26289. CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Li Y, Zhou M, Hu Q, Bai XC, Huang W, Scheres SH, Shi Y (2017) Mechanistic insights into caspase-9 activation by the structure of the apoptosome holoenzyme. Proc Natl Acad Sci U S A 114(7):1542–1547. CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Childers MC, Daggett V (2017) Insights from molecular dynamics simulations for computational protein design. Mol Syst Des Eng 2(1):9–33. CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Druskovic M, Suput D, Milisav I (2006) Overexpression of caspase-9 triggers its activation and apoptosis in vitro. Croat Med J 47(6):832–840PubMedPubMedCentralGoogle Scholar
  9. 9.
    Meszaros B, Erdos G, Dosztanyi Z (2018) IUPred2A: context-dependent prediction of protein disorder as a function of redox state and protein binding. Nucleic Acids Res 46(W1):W329–w337. CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Dosztanyi Z, Meszaros B, Simon I (2009) ANCHOR: web server for predicting protein binding regions in disordered proteins. Bioinformatics 25(20):2745–2746. CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Waterhouse A, Bertoni M, Bienert S, Studer G, Tauriello G, Gumienny R, Heer FT, de Beer TAP, Rempfer C, Bordoli L, Lepore R, Schwede T (2018) SWISS-MODEL: homology modelling of protein structures and complexes. Nucleic Acids Res 46(W1):W296–W303. CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Song Y, DiMaio F, Wang RY-R, Kim D, Miles C, Brunette T, Thompson J, Baker D (2013) High-resolution comparative modeling with RosettaCM. Structure 21(10):1735–1742. CrossRefPubMedGoogle Scholar
  13. 13.
    Xu D, Zhang Y (2012) Ab initio protein structure assembly using continuous structure fragments and optimized knowledge-based force field. Proteins 80(7):1715–1735. CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Kelley LA, Mezulis S, Yates CM, Wass MN, Sternberg MJE (2015) The Phyre2 web portal for protein modeling, prediction and analysis. Nat Protoc 10:845. CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Yang J, Yan R, Roy A, Xu D, Poisson J, Zhang Y (2015) The I-TASSER Suite: protein structure and function prediction. Nat Methods 12(1):7–8. CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Kallberg M, Wang H, Wang S, Peng J, Wang Z, Lu H, Xu J (2012) Template-based protein structure modeling using the RaptorX web server. Nat Protoc 7(8):1511–1522. CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Lovell SC, Davis IW, Arendall 3rd WB, de Bakker PI, Word JM, Prisant MG, Richardson JS, Richardson DC (2003) Structure validation by Calpha geometry: phi,psi and Cbeta deviation. Proteins 50(3):437–450. CrossRefPubMedGoogle Scholar
  18. 18.
    Chen VB, Arendall 3rd WB, Headd JJ, Keedy DA, Immormino RM, Kapral GJ, Murray LW, Richardson JS, Richardson DC (2010) MolProbity: all-atom structure validation for macromolecular crystallography. Acta Crystallogr D Biol Crystallogr 66(Pt 1):12–21. CrossRefPubMedGoogle Scholar
  19. 19.
    Laskowski RA, MacArthur MW, Moss DS, Thornton JM (1993) PROCHECK: a program to check the stereochemical quality of protein structures. J Appl Crystallogr 26(2):283–291CrossRefGoogle Scholar
  20. 20.
    Abraham MJ, Murtola T, Schulz R, Páll S, Smith JC, Hess B, Lindahl E (2015) GROMACS: High performance molecular simulations through multi-level parallelism from laptops to supercomputers. SoftwareX:Medium: ED; Size: p. 19-25. CrossRefGoogle Scholar
  21. 21.
    Jorgensen WL, Tirado-Rives J (1988) The OPLS [optimized potentials for liquid simulations] potential functions for proteins, energy minimizations for crystals of cyclic peptides and crambin. J Am Chem Soc 110(6):1657–1666. CrossRefPubMedGoogle Scholar
  22. 22.
    Ferguson DM (1995) Parameterization and evaluation of a flexible water model. J Comput Chem 16(4):501–511. CrossRefGoogle Scholar
  23. 23.
    Essmann U, Perera L, Berkowitz ML, Darden T, Lee H, Pedersen LG (1995) A smooth particle mesh Ewald method. J Chem Phys 103(19):8577–8593. CrossRefGoogle Scholar
  24. 24.
    Nosé S (1984) A molecular dynamics method for simulations in the canonical ensemble. Mol Phys 52(2):255–268. CrossRefGoogle Scholar
  25. 25.
    Hoover WG (1985) Canonical dynamics: Equilibrium phase-space distributions. Phys Rev A 31(3):1695–1697. CrossRefGoogle Scholar
  26. 26.
    Parrinello M, Rahman A (1981) Polymorphic transitions in single crystals: A new molecular dynamics method. J Appl Phys 52(12):7182–7190. CrossRefGoogle Scholar
  27. 27.
    DeLano WL (2002) Pymol: An open-source molecular graphics tool. CCP4 Newsletter On Protein Crystallography 40:82–92Google Scholar
  28. 28.
    Humphrey W, Dalke A, Schulten K (1996) VMD: Visual molecular dynamics. J Mol Graph 14(1):33–38. CrossRefPubMedGoogle Scholar
  29. 29.
    Shiozaki EN, Chai J, Shi Y (2002) Oligomerization and activation of caspase-9, induced by Apaf-1 CARD. Proc Natl Acad Sci U S A 99(7):4197–4202. CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    Huber KL, Serrano BP, Hardy JA (2018) Caspase-9 CARD: core domain interactions require a properly formed active site. Biochem J 475(6):1177–1196. CrossRefPubMedPubMedCentralGoogle Scholar
  31. 31.
    Hu Q, Wu D, Chen W, Yan Z, Yan C, He T, Liang Q, Shi Y (2014) Molecular determinants of caspase-9 activation by the Apaf-1 apoptosome. Proc Natl Acad Sci U S A 111(46):16254–16261. CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2020

Authors and Affiliations

  1. 1.Laboratorio de Investigación Bioquímica y Biofísica Computacional, Doctorado en Ciencias en Biotecnología, ENMHInstituto Politécnico Nacional, Guillermo Massieu HelgueraMexico CityMéxico
  2. 2.Laboratorio Transdisciplinario de Investigación en Sistemas EvolutivosSEPI de la ESCOM del Instituto Politécnico NacionalMexico CityMéxico

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