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. 2000 Jul;79(1):51–65. doi: 10.1016/S0006-3495(00)76273-4

The key event in force-induced unfolding of Titin's immunoglobulin domains.

H Lu 1, K Schulten 1
PMCID: PMC1300915  PMID: 10866937

Abstract

Steered molecular dynamics simulation of force-induced titin immunoglobulin domain I27 unfolding led to the discovery of a significant potential energy barrier at an extension of approximately 14 A on the unfolding pathway that protects the domain against stretching. Previous simulations showed that this barrier is due to the concurrent breaking of six interstrand hydrogen bonds (H-bonds) between beta-strands A' and G that is preceded by the breaking of two to three hydrogen bonds between strands A and B, the latter leading to an unfolding intermediate. The simulation results are supported by Angstrom-resolution atomic force microscopy data. Here we perform a structural and energetic analysis of the H-bonds breaking. It is confirmed that H-bonds between strands A and B break rapidly. However, the breaking of the H-bond between strands A' and G needs to be assisted by fluctuations of water molecules. In nanosecond simulations, water molecules are found to repeatedly interact with the protein backbone atoms, weakening individual interstrand H-bonds until all six A'-G H-bonds break simultaneously under the influence of external stretching forces. Only when those bonds are broken can the generic unfolding take place, which involves hydrophobic interactions of the protein core and exerts weaker resistance against stretching than the key event.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bryant Z., Pande V. S., Rokhsar D. S. Mechanical unfolding of a beta-hairpin using molecular dynamics. Biophys J. 2000 Feb;78(2):584–589. doi: 10.1016/S0006-3495(00)76618-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Carrion-Vazquez M., Marszalek P. E., Oberhauser A. F., Fernandez J. M. Atomic force microscopy captures length phenotypes in single proteins. Proc Natl Acad Sci U S A. 1999 Sep 28;96(20):11288–11292. doi: 10.1073/pnas.96.20.11288. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Carrion-Vazquez M., Oberhauser A. F., Fowler S. B., Marszalek P. E., Broedel S. E., Clarke J., Fernandez J. M. Mechanical and chemical unfolding of a single protein: a comparison. Proc Natl Acad Sci U S A. 1999 Mar 30;96(7):3694–3699. doi: 10.1073/pnas.96.7.3694. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Clarke J., Cota E., Fowler S. B., Hamill S. J. Folding studies of immunoglobulin-like beta-sandwich proteins suggest that they share a common folding pathway. Structure. 1999 Sep 15;7(9):1145–1153. doi: 10.1016/s0969-2126(99)80181-6. [DOI] [PubMed] [Google Scholar]
  5. Erickson H. P. Reversible unfolding of fibronectin type III and immunoglobulin domains provides the structural basis for stretch and elasticity of titin and fibronectin. Proc Natl Acad Sci U S A. 1994 Oct 11;91(21):10114–10118. doi: 10.1073/pnas.91.21.10114. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Evans E., Ritchie K. Strength of a weak bond connecting flexible polymer chains. Biophys J. 1999 May;76(5):2439–2447. doi: 10.1016/S0006-3495(99)77399-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Granzier H., Helmes M., Trombitás K. Nonuniform elasticity of titin in cardiac myocytes: a study using immunoelectron microscopy and cellular mechanics. Biophys J. 1996 Jan;70(1):430–442. doi: 10.1016/S0006-3495(96)79586-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Greaser M. L., Sebestyen M. G., Fritz J. D., Wolff J. A. cDNA sequence of rabbit cardiac titin/connectin. Adv Biophys. 1996;33:13–25. [PubMed] [Google Scholar]
  9. Grubmüller H., Heymann B., Tavan P. Ligand binding: molecular mechanics calculation of the streptavidin-biotin rupture force. Science. 1996 Feb 16;271(5251):997–999. doi: 10.1126/science.271.5251.997. [DOI] [PubMed] [Google Scholar]
  10. Helmes M., Trombitás K., Centner T., Kellermayer M., Labeit S., Linke W. A., Granzier H. Mechanically driven contour-length adjustment in rat cardiac titin's unique N2B sequence: titin is an adjustable spring. Circ Res. 1999 Jun 11;84(11):1339–1352. doi: 10.1161/01.res.84.11.1339. [DOI] [PubMed] [Google Scholar]
  11. Humphrey W., Dalke A., Schulten K. VMD: visual molecular dynamics. J Mol Graph. 1996 Feb;14(1):33-8, 27-8. doi: 10.1016/0263-7855(96)00018-5. [DOI] [PubMed] [Google Scholar]
  12. Isralewitz B., Izrailev S., Schulten K. Binding pathway of retinal to bacterio-opsin: a prediction by molecular dynamics simulations. Biophys J. 1997 Dec;73(6):2972–2979. doi: 10.1016/S0006-3495(97)78326-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Izrailev S., Stepaniants S., Balsera M., Oono Y., Schulten K. Molecular dynamics study of unbinding of the avidin-biotin complex. Biophys J. 1997 Apr;72(4):1568–1581. doi: 10.1016/S0006-3495(97)78804-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Kellermayer M. S., Granzier H. L. Elastic properties of single titin molecules made visible through fluorescent F-actin binding. Biochem Biophys Res Commun. 1996 Apr 25;221(3):491–497. doi: 10.1006/bbrc.1996.0624. [DOI] [PubMed] [Google Scholar]
  15. Kellermayer M. S., Smith S. B., Granzier H. L., Bustamante C. Folding-unfolding transitions in single titin molecules characterized with laser tweezers. Science. 1997 May 16;276(5315):1112–1116. doi: 10.1126/science.276.5315.1112. [DOI] [PubMed] [Google Scholar]
  16. Kosztin D., Izrailev S., Schulten K. Unbinding of retinoic acid from its receptor studied by steered molecular dynamics. Biophys J. 1999 Jan;76(1 Pt 1):188–197. doi: 10.1016/S0006-3495(99)77188-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Krammer A., Lu H., Isralewitz B., Schulten K., Vogel V. Forced unfolding of the fibronectin type III module reveals a tensile molecular recognition switch. Proc Natl Acad Sci U S A. 1999 Feb 16;96(4):1351–1356. doi: 10.1073/pnas.96.4.1351. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Labeit S., Kolmerer B., Linke W. A. The giant protein titin. Emerging roles in physiology and pathophysiology. Circ Res. 1997 Feb;80(2):290–294. doi: 10.1161/01.res.80.2.290. [DOI] [PubMed] [Google Scholar]
  19. Labeit S., Kolmerer B. Titins: giant proteins in charge of muscle ultrastructure and elasticity. Science. 1995 Oct 13;270(5234):293–296. doi: 10.1126/science.270.5234.293. [DOI] [PubMed] [Google Scholar]
  20. Lu H., Isralewitz B., Krammer A., Vogel V., Schulten K. Unfolding of titin immunoglobulin domains by steered molecular dynamics simulation. Biophys J. 1998 Aug;75(2):662–671. doi: 10.1016/S0006-3495(98)77556-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Lu H., Schulten K. Steered molecular dynamics simulations of force-induced protein domain unfolding. Proteins. 1999 Jun 1;35(4):453–463. [PubMed] [Google Scholar]
  22. Machado C., Sunkel C. E., Andrew D. J. Human autoantibodies reveal titin as a chromosomal protein. J Cell Biol. 1998 Apr 20;141(2):321–333. doi: 10.1083/jcb.141.2.321. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Marszalek P. E., Lu H., Li H., Carrion-Vazquez M., Oberhauser A. F., Schulten K., Fernandez J. M. Mechanical unfolding intermediates in titin modules. Nature. 1999 Nov 4;402(6757):100–103. doi: 10.1038/47083. [DOI] [PubMed] [Google Scholar]
  24. Maruyama K. Connectin/titin, giant elastic protein of muscle. FASEB J. 1997 Apr;11(5):341–345. doi: 10.1096/fasebj.11.5.9141500. [DOI] [PubMed] [Google Scholar]
  25. Nagar B., Overduin M., Ikura M., Rini J. M. Structural basis of calcium-induced E-cadherin rigidification and dimerization. Nature. 1996 Mar 28;380(6572):360–364. doi: 10.1038/380360a0. [DOI] [PubMed] [Google Scholar]
  26. Oberhauser A. F., Marszalek P. E., Erickson H. P., Fernandez J. M. The molecular elasticity of the extracellular matrix protein tenascin. Nature. 1998 May 14;393(6681):181–185. doi: 10.1038/30270. [DOI] [PubMed] [Google Scholar]
  27. Paci E., Karplus M. Forced unfolding of fibronectin type 3 modules: an analysis by biased molecular dynamics simulations. J Mol Biol. 1999 May 7;288(3):441–459. doi: 10.1006/jmbi.1999.2670. [DOI] [PubMed] [Google Scholar]
  28. Rief M., Gautel M., Oesterhelt F., Fernandez J. M., Gaub H. E. Reversible unfolding of individual titin immunoglobulin domains by AFM. Science. 1997 May 16;276(5315):1109–1112. doi: 10.1126/science.276.5315.1109. [DOI] [PubMed] [Google Scholar]
  29. Rief M., Gautel M., Schemmel A., Gaub H. E. The mechanical stability of immunoglobulin and fibronectin III domains in the muscle protein titin measured by atomic force microscopy. Biophys J. 1998 Dec;75(6):3008–3014. doi: 10.1016/S0006-3495(98)77741-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Rief M., Pascual J., Saraste M., Gaub H. E. Single molecule force spectroscopy of spectrin repeats: low unfolding forces in helix bundles. J Mol Biol. 1999 Feb 19;286(2):553–561. doi: 10.1006/jmbi.1998.2466. [DOI] [PubMed] [Google Scholar]
  31. Rohs R., Etchebest C., Lavery R. Unraveling proteins: a molecular mechanics study. Biophys J. 1999 May;76(5):2760–2768. doi: 10.1016/S0006-3495(99)77429-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Socci N. D., Onuchic J. N., Wolynes P. G. Stretching lattice models of protein folding. Proc Natl Acad Sci U S A. 1999 Mar 2;96(5):2031–2035. doi: 10.1073/pnas.96.5.2031. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Tskhovrebova L., Trinick J., Sleep J. A., Simmons R. M. Elasticity and unfolding of single molecules of the giant muscle protein titin. Nature. 1997 May 15;387(6630):308–312. doi: 10.1038/387308a0. [DOI] [PubMed] [Google Scholar]
  34. Wang K., McCarter R., Wright J., Beverly J., Ramirez-Mitchell R. Viscoelasticity of the sarcomere matrix of skeletal muscles. The titin-myosin composite filament is a dual-stage molecular spring. Biophys J. 1993 Apr;64(4):1161–1177. doi: 10.1016/S0006-3495(93)81482-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Wriggers W., Schulten K. Investigating a back door mechanism of actin phosphate release by steered molecular dynamics. Proteins. 1999 May 1;35(2):262–273. [PubMed] [Google Scholar]
  36. Yang G., Cecconi C., Baase W. A., Vetter I. R., Breyer W. A., Haack J. A., Matthews B. W., Dahlquist F. W., Bustamante C. Solid-state synthesis and mechanical unfolding of polymers of T4 lysozyme. Proc Natl Acad Sci U S A. 2000 Jan 4;97(1):139–144. doi: 10.1073/pnas.97.1.139. [DOI] [PMC free article] [PubMed] [Google Scholar]

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