Journal of Muscle Research and Cell Motility

, Volume 40, Issue 3–4, pp 389–398 | Cite as

CaATP prolongs strong actomyosin binding and promotes futile myosin stroke

  • Jinghua Ge
  • Akhil Gargey
  • Irina V. Nesmelova
  • Yuri E. NesmelovEmail author
Original Paper


Calcium plays an essential role in muscle contraction, regulating actomyosin interaction by binding troponin of thin filaments. There are several buffers for calcium in muscle, and those buffers play a crucial role in the formation of the transient calcium wave in sarcomere upon muscle activation. One such calcium buffer in muscle is ATP. ATP is a fuel molecule, and the important role of MgATP in muscle is to bind myosin and supply energy for the power stroke. Myosin is not a specific ATPase, and CaATP also supports myosin ATPase activity. The concentration of CaATP in sarcomeres reaches 1% of all ATP available. Since 294 myosin molecules form a thick filament, naïve estimation gives three heads per filament with CaATP bound, instead of MgATP. We found that CaATP dissociates actomyosin slower than MgATP, thus increasing the time of the strong actomyosin binding. The rate of the basal CaATPase is faster than that of MgATPase, myosin readily produces futile stroke with CaATP. When calcium is upregulated, as in malignant hyperthermia, kinetics of myosin and actomyosin interaction with CaATP suggest that myosin CaATPase activity may contribute to observed muscle rigidity and enhanced muscle thermogenesis.


Myosin Muscle ATP Calcium Malignant hyperthermia Transient kinetics 



This work was supported by National Institutes of Health (Grant No. HL132315) and by funds provided by the University of North Carolina at Charlotte.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflicts of interest with the contents of this article.

Ethical approval

Myosin and actin were produced from rabbit skeletal tissue. All experimental protocols were approved by the Institutional Animal Care and Use Committee of UNC Charlotte and all experiments were performed in accordance with relevant guidelines and regulations.

Supplementary material

10974_2019_9556_MOESM1_ESM.docx (120 kb)
Supplementary material 1 (DOCX 120 kb)


  1. Ali SZ, Taguchi A, Rosenberg H (2003) Malignant hyperthermia. Best Pract Res Clin Anaesthesiol 17:519–533PubMedGoogle Scholar
  2. Bagshaw CR (1975) Kinetic mechanism of manganous ion-dependent adenosine-triphosphatase of myosin subfragment 1. FEBS Lett 58:197–201. CrossRefPubMedGoogle Scholar
  3. Bagshaw CR, Trentham DR (1973) The reversibility of adenosine triphosphate cleavage by myosin. Biochem J 133:323–328PubMedPubMedCentralGoogle Scholar
  4. Bagshaw CR, Trentham DR (1974) The characterization of myosin-product complexes and of product-release steps during the magnesium ion-dependent adenosine triphosphatase reaction. Biochem J 141:331–349PubMedPubMedCentralGoogle Scholar
  5. Banerjee S, Morkin E (1978) Thermodynamic studies on the binding of adenosine diphosphate and calcium to beef cardiac myosin. Biochim Biophys Acta 536:10–17PubMedGoogle Scholar
  6. Baylor SM, Hollingworth S (1998) Model of sarcomeric Ca2+ movements, including ATP Ca2+ binding and diffusion, during activation of frog skeletal muscle. J Gen Physiol 112:297–316. CrossRefPubMedPubMedCentralGoogle Scholar
  7. Baylor SM, Hollingworth S (2012) Intracellular calcium movements during excitation-contraction coupling in mammalian slow-twitch and fast-twitch muscle fibers. J Gen Physiol 139:261–272. CrossRefPubMedPubMedCentralGoogle Scholar
  8. Craig R, Offer G (1976) Axial arrangement of crossbridges in thick filaments of vertebrate skeletal muscle. J Mol Biol 102:325–332PubMedGoogle Scholar
  9. Criddle AH, Geeves MA, Jeffries T (1985) The use of actin labelled with N-(1-pyrenyl)iodoacetamide to study the interaction of actin with myosin subfragments and troponin/tropomyosin. Biochem J 232:343–349PubMedPubMedCentralGoogle Scholar
  10. Cully TR, Edwards JN, Launikonis BS (2014) Activation and propagation of Ca2+ release from inside the sarcoplasmic reticulum network of mammalian skeletal muscle. J Physiol 592:3727–3746PubMedPubMedCentralGoogle Scholar
  11. Dancker P, Low I, Hasselbach W, Wieland T (1975) Interaction of actin with phalloidin - polymerization and stabilization of F-actin. Biochim Biophys Acta 400:407–414. CrossRefPubMedGoogle Scholar
  12. De La Cruz EM, Ostap EM (2009) Kinetic and equilibrium analysis of the myosin ATPase. Methods Enzymol 455:157–192Google Scholar
  13. De La Cruz EM, Wells AL, Sweeney HL, Ostap EM (2000) Actin and light chain isoform dependence of myosin V kinetics. Biochemistry-US 39:14196–14202Google Scholar
  14. Deacon JC, Bloemink MJ, Rezavandi H, Geeves MA, Leinwand LA (2012) Erratum to: Identification of functional differences between recombinant human alpha and beta cardiac myosin motors. Cell Mol Life Sci 69:4239–4255. CrossRefPubMedPubMedCentralGoogle Scholar
  15. Finer JT, Simmons RM, Spudich JA (1994) Single myosin molecule mechanics: piconewton forces and nanometre steps. Nature 368:113–119PubMedGoogle Scholar
  16. Fusi L et al (2017) Minimum number of myosin motors accounting for shortening velocity under zero load in skeletal muscle. J Physiol 595:1127–1142PubMedGoogle Scholar
  17. Ge J, Bouriyaphone SD, Serebrennikova TA, Astashkin AV, Nesmelov YE (2016) Macromolecular crowding modulates actomyosin kinetics. Biophys J 111:178–184PubMedPubMedCentralGoogle Scholar
  18. Geeves MA (1989) Dynamic interaction between actin and myosin subfragment 1 in the presence of ADP. Biochemistry-US 28:5864–5871Google Scholar
  19. Greenberg MJ, Moore JR (2010) The molecular basis of frictional loads in the in vitro motility assay with applications to the study of the loaded mechanochemistry of molecular motors. Cytoskeleton (Hoboken) 67:273–285. CrossRefGoogle Scholar
  20. Guhathakurta P, Prochniewicz E, Roopnarine O, Rohde JA, Thomas DD (2017) A cardiomyopathy mutation in the myosin essential light chain alters actomyosin structure. Biophys J 113:91–100PubMedPubMedCentralGoogle Scholar
  21. Harada Y, Sakurada K, Aoki T, Thomas DD, Yanagida T (1990) Mechanochemical coupling in actomyosin energy transduction studied by in vitro movement assay. J Mol Biol 216:49–68PubMedGoogle Scholar
  22. Harris DE, Warshaw DM (1993) Smooth and skeletal muscle myosin both exhibit low duty cycles at zero load in vitro. J Biol Chem 268:14764–14768PubMedGoogle Scholar
  23. Heissler SM, Liu X, Korn ED, Sellers JR (2013) Kinetic characterization of the ATPase and actin-activated ATPase activities of Acanthamoeba castellanii myosin-2. J Biol Chem 288:26709–26720PubMedPubMedCentralGoogle Scholar
  24. Henn A, De La Cruz EM (2005) Vertebrate myosin VIIb is a high duty ratio motor adapted for generating and maintaining tension. J Biol Chem 280:39665–39676PubMedGoogle Scholar
  25. Hooijman P, Stewart MA, Cooke R (2011) A new state of cardiac myosin with very slow ATP turnover: a potential cardioprotective mechanism in the heart. Biophys J 100:1969–1976. CrossRefPubMedPubMedCentralGoogle Scholar
  26. Houk TW Jr, Ue K (1974) The measurement of actin concentration in solution: a comparison of methods. Anal Biochem 62:66–74PubMedGoogle Scholar
  27. Kawai M, Wray JS, Zhao Y (1993) The effect of lattice spacing change on cross-bridge kinetics in chemically skinned rabbit psoas muscle-fibers. 1. Proportionality between the lattice spacing and the fiber width. Biophys J 64:187–196. CrossRefPubMedPubMedCentralGoogle Scholar
  28. Lanzetta PA, Alvarez LJ, Reinach PS, Candia OA (1979) An improved assay for nanomole amounts of inorganic phosphate. Anal Biochem 100:95–97PubMedGoogle Scholar
  29. Lymn RW, Taylor EW (1971) Mechanism of adenosine triphosphate hydrolysis by actomyosin. Biochemistry-US 10:4617–4624. CrossRefGoogle Scholar
  30. MacLennan DH, Phillips MS (1992) Malignant hyperthermia. Science 256:789–794PubMedGoogle Scholar
  31. Manno C et al (2013) Altered Ca2+ concentration, permeability and buffering in the myofibre Ca2+ store of a mouse model of malignant hyperthermia. J Physiol 591:4439–4457. CrossRefPubMedPubMedCentralGoogle Scholar
  32. Margossian SS, Lowey S (1982) Preparation of myosin and its subfragments from rabbit skeletal muscle. Methods Enzymol 85:55–71PubMedGoogle Scholar
  33. Marston SB, Taylor EW (1980) Comparison of the myosin and actomyosin ATPase mechanisms of the four types of vertebrate muscles. J Mol Biol 139:573–600PubMedGoogle Scholar
  34. Martonosi A, Gouvea MA, Gergely J (1960) Studies on actin. 1. Interaction of C-14-labeled adenine nucleotides with actin. J Biol Chem 235:1700–1703PubMedGoogle Scholar
  35. Melchior NC (1954) Sodium and potassium complexes of adenosinetriphosphate: equilibrium studies. J Biol Chem 208:615–627PubMedGoogle Scholar
  36. Nyitrai M, Rossi R, Adamek N, Pellegrino MA, Bottinelli R, Geeves MA (2006) What limits the velocity of fast-skeletal muscle contraction in mammals? J Mol Biol 355:432–442. CrossRefPubMedGoogle Scholar
  37. Polosukhina K, Eden D, Chinn M, Highsmith S (2000) CaATP as a substrate to investigate the myosin lever arm hypothesis of force generation. Biophys J 78:1474–1481PubMedPubMedCentralGoogle Scholar
  38. Rosenberg H, Davis M, James D, Pollock N, Stowell K (2007) Malignant hyperthermia. Orphanet J Rare Dis 2:21. CrossRefPubMedPubMedCentralGoogle Scholar
  39. Rosenfeld SS, Taylor EW (1984) The ATPase mechanism of skeletal and smooth muscle acto-subfragment 1. J Biol Chem 259:11908–11919PubMedGoogle Scholar
  40. Rosenfeld SS, Taylor EW (1987) The mechanism of regulation of actomyosin subfragment-1 ATPase. J Biol Chem 262:9984–9993PubMedGoogle Scholar
  41. Rosenfeld SS et al (2000) Kinetic and spectroscopic evidence for three actomyosin: ADP states in smooth muscle. J Biol Chem 275:25418–25426. CrossRefPubMedGoogle Scholar
  42. Segel IH (1976) Biochemical calculations. Wiley, New YorkGoogle Scholar
  43. Shriver JW, Sykes BD (1981) Phosphorus-31 nuclear magnetic resonance evidence for two conformations of myosin subfragment-1. Nucleotide complexes. Biochemistry-US 20:2004–2012Google Scholar
  44. Siemankowski RF, Wiseman MO, White HD (1985) ADP dissociation from actomyosin subfragment 1 is sufficiently slow to limit the unloaded shortening velocity in vertebrate muscle. Proc Natl Acad Sci USA 82:658–662PubMedGoogle Scholar
  45. Sigel H, Song B (1996) Solution studies of nucleotide-metal ion complexes - isomeric equilibria. In: Astrid Sigel HS (ed) Metal ions in biological systems, vol 32. CRC Press, Boca Raton, p 135Google Scholar
  46. Sleep JA, Trybus KM, Johnson KA, Taylor EW (1981) Kinetic-studies of normal and modified heavy-meromyosin and subfragment. 1. J Muscle Res Cell Motil 2:373–399. CrossRefGoogle Scholar
  47. Sommese RF et al (2013) Molecular consequences of the R453C hypertrophic cardiomyopathy mutation on human beta-cardiac myosin motor function. Proc Natl Acad Sci USA 110:12607–12612PubMedGoogle Scholar
  48. Struk A, Lehmann-Horn F, Melzer W (1998) Voltage-dependent calcium release in human malignant hyperthermia muscle fibers. Biophys J 75:2402–2410PubMedPubMedCentralGoogle Scholar
  49. Strzelecka-Golaszewska H, Prochniewicz E, Nowak E, Zmorzynski S, Drabikowski W (1980) Chicken-gizzard actin: polymerization and stability. Eur J Biochem 104:41–52PubMedGoogle Scholar
  50. Takagi Y, Yang Y, Fujiwara I, Jacobs D, Cheney RE, Sellers JR, Kovacs M (2008) Human myosin Vc is a low duty ratio, nonprocessive molecular motor. J Biol Chem 283:8527–8537. CrossRefPubMedGoogle Scholar
  51. Tkachev YV, Ge J, Negrashov IV, Nesmelov YE (2013) Metal cation controls myosin and actomyosin kinetics. Protein Sci 22:1766–1774PubMedPubMedCentralGoogle Scholar
  52. Travers F, Hillaire D (1979) Cryoenzymological studies on myosin subfragment 1 solvent, temperature and pH effects on the overall reaction. Eur J Biochem 98:293–299PubMedGoogle Scholar
  53. Uyeda TQ, Kron SJ, Spudich JA (1990) Myosin step size. Estimation from slow sliding movement of actin over low densities of heavy meromyosin. J Mol Biol 214:699–710PubMedGoogle Scholar
  54. Waller GS, Ouyang G, Swafford J, Vibert P, Lowey S (1995) A minimal motor domain from chicken skeletal muscle myosin. J Biol Chem 270:15348–15352PubMedGoogle Scholar
  55. Webb M, Jackson DR Jr, Stewart TJ, Dugan SP, Carter MS, Cremo CR, Baker JE (2013) The myosin duty ratio tunes the calcium sensitivity and cooperative activation of the thin filament. Biochemistry-US 52:6437–6444Google Scholar
  56. White HD, Belknap B, Jiang W (1993) Kinetics of binding and hydrolysis of a series of nucleoside triphosphates by actomyosin-S1. Relationship between solution rate constants and properties of muscle fibers. J Biol Chem 268:10039–10045PubMedGoogle Scholar
  57. Woodward SKA, Eccleston JF, Geeves MA (1991) Kinetics of the interaction of 2′(3′)-O-(N-methylanthraniloyl)-ATP with myosin subfragment-1 and actomyosin subfragment-1- characterization of 2 Acto. S1.Adp complexes. Biochemistry-US 30:422–430. CrossRefGoogle Scholar
  58. Yengo CM, Takagi Y, Sellers JR (2012) Temperature dependent measurements reveal similarities between muscle and non-muscle myosin motility. J Muscle Res Cell M 33:385–394Google Scholar

Copyright information

© Springer Nature Switzerland AG 2019

Authors and Affiliations

  • Jinghua Ge
    • 1
    • 2
  • Akhil Gargey
    • 1
    • 2
    • 3
  • Irina V. Nesmelova
    • 1
    • 2
  • Yuri E. Nesmelov
    • 1
    • 2
    Email author
  1. 1.Department of Physics and Optical ScienceUniversity of North Carolina CharlotteCharlotteUSA
  2. 2.Center for Biomedical Engineering and ScienceUniversity of North CarolinaCharlotteUSA
  3. 3.Department of Biological ScienceUniversity of North Carolina CharlotteCharlotteUSA

Personalised recommendations