Abstract
Through the study of single molecules it has become possible to explain the function of many of the complex molecular assemblies found in cells1,2,3,4,5. The protein titin provides muscle with its passive elasticity. Each titin molecule extends over half a sarcomere, and its extensibility has been studied both in situ6,7,8,9,10 and at the level of single molecules11,12,13,14. These studies suggested that titin is not a simple entropic spring but has a complex structure-dependent elasticity. Here we use protein engineering and single-molecule atomic force microscopy15 to examine the mechanical components that form the elastic region of human cardiac titin16,17. We show that when these mechanical elements are combined, they explain the macroscopic behaviour of titin in intact muscle6. Our studies show the functional reconstitution of a protein from the sum of its parts.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 51 print issues and online access
$199.00 per year
only $3.90 per issue
Rent or buy this article
Prices vary by article type
from$1.95
to$39.95
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
References
Sigworth, F. J. & Neher, E. Single Na+ channel currents observed in cultured rat muscle cells. Nature 287, 447–449 (1980)
Bustamante, C., Smith, S. B., Liphardt, J. & Smith, D. Single-molecule studies of DNA mechanics. Curr. Opin. Struct. Biol. 10, 279–285 (2000)
Smith, D. E. et al. The bacteriophage φ29 portal motor can package DNA against a large internal force. Nature 413, 748–752 (2001)
Finer, J. T., Simmons, R. M. & Spudich, J. A. Single myosin molecule mechanics: piconewton forces and nanometre steps. Nature 368, 113–119 (1994)
Lu, H. & Schulten, K. Steered molecular dynamics simulations of force-induced protein domain unfolding. Proteins Struct. Funct. Genet. 35, 453–463 (1999)
Linke, W. A. et al. I-band titin in cardiac muscle is a three-element molecular spring and is critical for maintaining thin filament structure. J. Cell Biol. 146, 631–644 (1999)
Maruyama, K. Connectin/titin, giant elastic protein of muscle. FASEB J. 11, 341–345 (1997)
Wang, K. Titin/connectin and nebulin: giant protein rulers of muscle structure and function. Adv. Biophys. 33, 123–134 (1996)
Gregorio, C. C., Granzier, H., Sorimachi, H. & Labeit, S. Muscle assembly: a titanic achievement? Curr. Opin. Cell Biol. 11, 18–25 (1999)
Trinick, J. & Tskhovrebova, L. Titin: a molecular control freak. Trends Cell Biol. 9, 377–380 (1999)
Rief, M., Gautel, M., Oesterhelt, F., Fernandez, J. M. & Gaub, H. E. Reversible unfolding of individual immunoglobin domains by AFM. Science 276, 1109–1112 (1997)
Kellermayer, M., Smith, S., Granzier, H. & Bustamante, C. Folding-unfolding transitions in single titin molecules characterized with laser tweezers. Science 276, 1112–1116 (1997)
Tskhovrebova, L., Trinick, J., Sleep, J. A. & Simmons, R. M. Elasticity and unfolding of single molecules of the giant muscle protein titin. Nature 387, 308–312 (1997)
Tskhovrebova, L. & Trinick, J. Direct visualization of extensibility in isolated titin molecules. J. Mol. Biol. 265, 100–106 (1997)
Fisher, T. E., Marszalek, P. E. & Fernandez, J. M. Stretching single molecules into novel conformations using the atomic force microscope. Nature Struct. Biol. 7, 719–724 (2000)
Freiburg, A. et al. Series of exon-skipping events in the elastic spring region of titin as the structural basis for myofibrillar elastic diversity. Circ. Res. 86, 1114–1121 (2000)
Labeit, S. & Kolmerer, B. Titins, giant proteins in charge of muscle ultrastructure and elasticity. Science 270, 293–296 (1995)
Carrion-Vazquez, M. et al. Mechanical and chemical unfolding of a single protein: a comparison. Proc. Natl Acad. Sci. USA 96, 3694–3699 (1999)
Li, H. B., Oberhauser, A. F., Fowler, S. B., Clarke, J. & Fernandez, J. M. Atomic force microscopy reveals the mechanical design of a modular protein. Proc. Natl Acad. Sci. USA 97, 6527–6531 (2000)
Li, H. B. et al. Multiple conformations of PEVK proteins detected by single-molecule techniques. Proc. Natl Acad. Sci. USA 98, 10682–10686 (2001)
Best, R. B., Li, B., Steward, A., Daggett, V. & Clarke, J. Can non-mechanical proteins withstand force? Stretching barnase by atomic force microscopy and molecular dynamics simulation. Biophys. J. 81, 2344–2356 (2001)
Marko, J. F. & Siggia, E. D. Stretching DNA. Macromolecules 28, 8759–8770 (1995)
Bell, G. I. Models for the specific adhesion of cells to cells. Science 200, 618–627 (1978)
Marszalek, P. E. et al. Mechanical unfolding intermediates in titin modules. Nature 402, 100–103 (1999)
Trombitas, K., Freiburg, A., Centner, T., Labeit, S. & Granzier, H. Molecular dissection of N2B cardiac titin's extensibility. Biophys. J. 77, 3189–3196 (1999)
Bueche, F. Physical Properties of Polymers 37 (Interscience, New York, 1962)
Linke, W. A. et al. Towards a molecular understanding of the elasticity of titin. J. Mol. Biol. 261, 62–71 (1996)
Higuchi, H., Nakauchi, Y., Maruyama, K. & Fujime, S. Characterization of beta-connectin (titin 2) from striated muscle by dynamic light scattering. Biophys. J. 65, 1906–1915 (1993)
Allen, D. G. & Kentish, J. C. The cellular basis of the length-tension relation in cardiac muscle. J. Mol. Cell Cardiol. 17, 821–840 (1985)
Scott, K. A., Steward, A., Fowler, S. B. & Clarke, J. Titin; a multidomain protein that behaves as the sum of its parts. J. Mol. Biol. 315, 819–829 (2002)
Acknowledgements
We thank H. Erickson for the electron microscope pictures of I27 polyproteins. W.A.L. thanks the German Research Foundation for a Heisenberg fellowship. This work was supported by the National Institutes of Health (J.M.F.)
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Competing interests
The authors declare that they have no competing financial interests.
Supplementary information
Rights and permissions
About this article
Cite this article
Li, H., Linke, W., Oberhauser, A. et al. Reverse engineering of the giant muscle protein titin. Nature 418, 998–1002 (2002). https://doi.org/10.1038/nature00938
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1038/nature00938
This article is cited by
-
Structural heterogeneity of the ion and lipid channel TMEM16F
Nature Communications (2024)
-
Cartilage-like protein hydrogels engineered via entanglement
Nature (2023)
-
The role of single-protein elasticity in mechanobiology
Nature Reviews Materials (2022)
-
Intramolecular hydrogen bonds in a single macromolecule: Strength in high vacuum versus liquid environments
Nano Research (2022)
-
Protein nanomechanics in biological context
Biophysical Reviews (2021)
Comments
By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.