13.1. Abstruct
Tropomyosin (Tm) is a 400 Å long coiled coil protein, and with troponin it regulates contraction in skeletal and cardiac muscles in a [Ca2+]-dependent manner. Tm consists of multiple domains with diverse stabilities in the coiled coil form, thus providing Tm with dynamic flexibility. This flexibility must play important roles in the actin binding and the cooperative transition between the calcium regulated states of the entire muscle thin filament. In order to understand the flexibility of Tm in its entirety, the atomic coordinates of Tm are needed. Here we report the two crystal structures of Tm segments. One is rabbit skeletal muscle α-Tm encompassing residues 176–284 with an N-terminal extension of 25 residues from the leucine zipper sequence of GCN4, which includes the region that interacts with the troponin core domain. The other is α-Tm encompassing residues 176–273 with N- and C-terminal extensions of the leucine zipper sequences. These two crystal structures imply that this molecule is a flexible coiled coil. First, Tm’s are not homogeneous and smooth coiled coils, but instead they undulate, with highly fluctuating local parameters specifying the coiled coil. Independent fluctuating showed by two crystal structures is important. Second, in the first crystal, the coiled coil is bent by 9 degrees in the region centered about Y214-E218-Y221, where the inter-helical distance has its maximum. On the other hand, no bend is observed at the same region in the second crystal even if its inter-helical distance has also its maximum. E218, an unusual negatively charged residue at the a position in the heptad repeat, seems to play the key role in destabilizing the coiled coil with alanine destabilizing clusters.
This is a preview of subscription content, log in via an institution.
Buying options
Tax calculation will be finalised at checkout
Purchases are for personal use only
Learn about institutional subscriptionsPreview
Unable to display preview. Download preview PDF.
13.7. References
K. Bailey, Tropomyosin: a new asymmetric protein component of muscle, Nature 157, 368–369 (1946).
S. Ebashi, Calcium ions and muscle contraction, Nature 240(5378), 217–218 (1972).
S. Ebashi, and M. Endo, Calcium ion and muscle contraction, Prog. Biophys. Mol. Biol. 18, 123–183 (1968).
S. V. Perry, Vertebrate tropomyosin: distribution, properties and function, J. Muscle Res. Cell Motil. 22(1), 5–49 (2001).
J. P. Lees-Miller, and D. M. Helfman, The molecular basis for tropomyosin isoform diversity, Bioessays 13(9), 429–437 (1991).
I. Ohtsuki, K. Maruyama, and S. Ebashi, Regulatory and cytoskeletal proteins of vertebrate skeletal muscle, Adv. Protein. Chem. 38, 1–67 (1986).
A. N. Lupas, and M. Gruber, The structure of alpha-helical coiled coils, Adv. Protein. Chem. 70, 37–78 (2005).
F. H. Crick, Is alpha-keratin a coiled coil?, Nature 170(4334), 882–883 (1952).
T. Alber, Structure of the leucine zipper, Curr. Opin. Genet. Dev. 2(2), 205–210 (1992).
D. N. Marti, and H. R. Bosshard, Electrostatic interactions in leucine zippers: thermodynamic analysis of the contributions of Glu and His residues and the effect of mutating salt bridges, J. Mol. Biol. 330(3), 621–637 (2003).
B. Tripet, K. Wagschal, P. Lavigne, C. T. Mant, and R. S. Hodges, Effects of side-chain characteristics on stability and oligomerization state of a de novo-designed model coiled-coil: 20 amino acid substitutions in position “d”, J. Mol. Biol. 300(2), 377–402 (2000).
K. Wagschal, B. Tripet, P. Lavigne, C. Mant, and R. S. Hodges, The role of position a in determining the stability and oligomerization state of alpha-helical coiled coils: 20 amino acid stability coefficients in the hydrophobic core of proteins, Protein Sci. 8(11), 2312–2329 (1999).
E. K. O’Shea, J. D. Klemm, P. S. Kim, and T. Alber, X-ray structure of the GCN4 leucine zipper, a two-stranded, parallel coiled coil, Science 254(5031), 539–544 (1991).
S. M. Lu, and R. S. Hodges, Defining the minimum size of a hydrophobic cluster in two-stranded alpha-helical coiled-coils: effects on protein stability, Protein Sci. 13(3), 714–726 (2004).
S. C. Kwok, and R. S. Hodges, Stabilizing and destabilizing clusters in the hydrophobic core of long two-stranded alpha-helical coiled-coils, J. Biol. Chem. 279(20), 21576–21588 (2004).
A. Singh, and S. E. Hitchcock-DeGregori, Local destabilization of the tropomyosin coiled coil gives the molecular flexibility required for actin binding, Biochemistry 42(48), 14114–14121 (2003).
K. I. Sano, K. Maeda, H. Taniguchi, and Y. Maeda, Amino-acid replacements in an internal region of tropomyosin alter the properties of the entire molecule, Eur. J. Biochem. 267(15), 4870–4877 (2000).
E. F. Woods, The conformational stabilities of tropomyosins, Aust. J. Biol. Sci. 29(5–6), 405–418 (1976).
A. Sato, and K. Mihashi, Thermal modification of structure of tropomyosin. I. Changes in the intensity and polarization of the intrinsic fluorescence (tyrosine), J. Biochem. (Tokyo) 71(4), 597–605 (1972).
S. S. Lehrer, Effects of an interchain disulfide bond on tropomyosin structure: intrinsic fluorescence and circular dichroism studies, J. Mol. Biol. 118(2), 209–226 (1978).
P. Graceffa, and S. S. Lehrer, The excimer fluorescence of pyrene-labeled tropomyosin. A probe of conformational dynamics, J. Biol. Chem. 255(23), 11296–11300 (1980).
S. L. Betcher-Lange, and S. S. Lehrer, Pyrene excimer fluorescence in rabbit skeletal alphaalphatropo-myosin labeled with N-(1-pyrene)maleimide. A probe of sulfhydryl proximity and local chain separation, J. Biol. Chem. 253(11), 3757–3760 (1978).
D. R. Betteridge, and S. S. Lehrer, Two conformational states of didansylcystine-labeled rabbit cardiac tropomyosin, J. Mol. Biol. 167(2), 481–496 (1983).
P. Graceffa, and S. S. Lehrer, Dynamic equilibrium between the two conformational states of spin-labeled tropomyosin., Biochemistry 23(12), 2606–2612 (1984).
B. F. Edwards, and B. D. Sykes, Nuclear magnetic resonance evidence for the coexistence of several conformational states of rabbit cardiac and skeletal tropomyosins, Biochemistry 19(12), 2577–2583 (1980).
S. A. Potekhin, and P. L. Privalov, Co-operative blocks in tropomyosin, J. Mol. Biol. 159(3), 519–535 (1982).
D. L. Williams Jr., and C. A. Swenson, Tropomyosin stability: assignment of thermally induced conformational transitions to separate regions of the molecule, Biochemistry 20(13), 3856–3864 (1981).
F. G. Whitby, and G. N. Phillips Jr., Crystal structure of tropomyosin at 7 Angstroms resolution, Proteins 38(1), 49–59 (2000).
J. H. Brown, K. H. Kim, G. Jun, N. J. Greenfield, R. Dominguez, N. Volkmann, S. E. Hitchcock-DeGregori, and C. Cohen, Deciphering the design of the tropomyosin molecule, Proc. Natl. Acad. Sci. USA 98(15), 8496–8501 (2001).
Li, Y., S. Mui, J. H. Brown, J. Strand, L. Reshetnikova, L. S. Tobacman, and C. Cohen, The crystal structure of the C-terminal fragment of striated-muscle alpha-tropomyosin reveals a key troponin T recognition site, Proc. Natl. Acad. Sci. USA 99(11), 7378–7383 (2002).
N. J. Greenfield, G. V. Swapna, Y. Huang, T. Palm, S. Graboski, G. T. Montelione, and S. E. Hitchcock-DeGregori, The structure of the carboxyl terminus of striated alpha-tropomyosin in solution reveals an unusual parallel arrangement of interacting alpha-helices, Biochemistry 42(3), 614–619 (2003).
N. Ookubo, Intramolecular disulfide linked alphabeta and alphaalpha in oxidized tropomyosin: separation, identification, and process of formation, J. Biochem. (Tokyo) 81(4), 923–931 (1977).
T. Shimizu, K. Ihara, R. Maesaki, M. Amano, K. Kaibuchi, and T. Hakoshima, Parallel coiled-coil association of the RhoA-binding domain in Rho-kinase, J. Biol. Chem. 278(46), 46046–46051 (2003).
M. V. Vinogradova, D. B. Stone, G. G. Malanina, C. Karatzaferi, R. Cooke, R. A. Mendelson, and R. J. Fletterick, Ca(2+)-regulated structural changes in troponin, Proc. Natl. Acad. Sci. USA 102(14), 5038–5043 (2005).
S. Takeda, A. Yamashita, K. Maeda, and Y. Maeda, Structure of the core domain of human cardiac troponin in the Ca(2+)-saturated form, Nature 424(6944), 35–41 (2003).
R. Maytum, F. Bathe, M. Konrad, and M. A. Geeves, Tropomyosin exon 6b is troponin-specific and required for correct acto-myosin regulation, J. Biol. Chem. 279(18), 18203–18209 (2004).
L. Kluwe, K. Maeda, A. Miegel, S. Fujita-Becker, Y. Maeda, G. Talbo, T. Houthaeve, and R. Kellner, Rabbit skeletal muscle alpha alpha-tropomyosin expressed in baculovirus-infected insect cells possesses the authentic N-terminus structure and functions, J. Muscle Res. Cell Motil. 16(2), 103–110 (1995).
M. Sugahara, and M. Miyano, Development of high-throughput automatic protein crystallization and observation system, Tanpakushitsu Kakusan Koso 47(8 Suppl), 1026–1032 (2002).
S. Adachi, T. Oguchi, H. Tanida, S.-Y. Park, H. Shimizu, H. Miyatake, N. Kamiya, Y. Shiro, Y. Inoue, T. Ueki, and T. Iizuka, The RIKEN Structural Biology Beamline II (BL44B2) at the SPring-8, Nucl. Instrum. Methods Phys. Res. A 467, 711–714 (2001).
J. W. Pflugrath, The finer things in X-ray diffraction data collection, Acta Crystallogr. D Biol. Crystallogr. 55(Pt 10), 1718–1725 (1999).
J. Navaza, Implementation of molecular replacement in AMoRe, Acta Crystallogr. D Biol. Crystallogr. 57(Pt 10), 1367–1372 (2001).
E. Potterton, P. Briggs, M. Turkenburg, and E. Dodson, A graphical user interface to the CCP4 program suite, Acta Crystallogr. D Biol. Crystallogr. 59(Pt 7), 1131–1137 (2003).
Collaborative Computational Project, Number 4, The CCP4 suite: programs for protein crystallography, Acta Crystallogr. D Biol. Crystallogr. 50(Pt 5), 760–763 (1994).
D. E. McRee, XtalView/Xfit — A versatile program for manipulating atomic coordinates and electron density, J. Struct. Biol. 125(2–5), 156–165 (1999).
D. E. McRee, Differential evolution for protein crystallographic optimizations, Acta Crystallogr. D Biol. Crystallogr. 60 (Pt 12, No 1), 2276–2279 (2004).
S. V. Strelkov, and P. Burkhard, Analysis of alpha-helical coiled coils with the program TWISTER reveals a structural mechanism for stutter compensation, J. Struct. Biol. 137(1–2), 54–64 (2002).
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2007 Springer
About this paper
Cite this paper
Nitanai, Y., Minakata, S., Maeda, K., Oda, N., Maéda, Y. (2007). Crystal Structures of Tropomyosin: Flexible Coiled-Coil. In: Ebashi, S., Ohtsuki, I. (eds) Regulatory Mechanisms of Striated Muscle Contraction. Advances in Experimental Medicine and Biology, vol 592. Springer, Tokyo. https://doi.org/10.1007/978-4-431-38453-3_13
Download citation
DOI: https://doi.org/10.1007/978-4-431-38453-3_13
Publisher Name: Springer, Tokyo
Print ISBN: 978-4-431-38451-9
Online ISBN: 978-4-431-38453-3
eBook Packages: Biomedical and Life SciencesBiomedical and Life Sciences (R0)