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So far the biggest actin binding protein not just in its size (600-900kDa) but in that it binds a potential 200 actin monomers! Each monomer is bound by a 35 amino acid residue that may also bind calmodulin, tropomyosin and troponin (Wang et al, 1996), and the N-terminal region binds tropomodulin (McElhinny et al, 2001). The actin-binding core of the repeat is around a conserved motif SDxxYK (Pfuhl et al, 1994), and this is proposed to bind actin through a cleft formed "behind" sub-domains 3 and 4  of actin in the standard view (Kabsch et al, 1990). Models have been proposed (Wang, 1999) in which it is suggested that nebulin co-aligns with tropomyosin and troponn to form a multifilament that wraps around the thin filament.  However, the model suggested by (Pfuhl et al, 1994) would seem to contradict this view as the proposed nebulin site is distinct from tropomysoin.  

Figure 1.  The Structure of Nebulin. 7x 35 amino-acid repeats form a unit that is itself repeated about 22 times.  The N-terminal associates with tropomodulin and so the pointed end of the thin filament the C-terminus binds components of the Z-disc.

The interaction of nebulin with actin is Ca2+-calmodulin sensitive (Root & Wang, 1994). Nebulin is proposed to form a "molecular ruler" controlling the length of the thin filament (Chen et al, 1993b), as differences in the length of expressed nebulin corresponds to the length of the sarcomere (Kruger et al, 1991; Labiet et al, 1991).  Nebulin is associated with an autosomal recessive nemaline myopathy in humans (Pelin et al, 1999).

Douglas Root Lab UCLA  Annalisa Pastore EMBL  Mark Pfuhl University of Leicester


Atsuta, F., Sato, K., Maruyama, K. & Shimada, Y. (1993) Distribution of connectin (titin), nebulin and a-actinin at myotendinous junctions of chicken pectoralis muscles: an immunofluorescence and immunoelectron microscopic study., J.Muscle Res. Cell Motil. 14, 511-517.

Bang, M. L., Mudry, R. E., McElhinny, A. S., Trombitas, K., Geach, A. J., Yamasaki, R., Sorimachi, H., Granzier, H., Gregorio, C. C. & Labeit, S. (2001) Myopalladin, a novel 145-kilodalton sarcomeric protein with mutliple roles in Z-disc and I-band assemblies., J.Cell Biol. 153, 413-427.

Chen, M.-J. G., Shih, C.-L. & Wang, K. (1993a) Nebulin as an actin zipper. A two-module nebulin fragment promotes actin nucleation and stabilizes actin filaments, J. Biol. Chem. 268, 20327-20334.

Chen, M.-J. G., Shih, C.-L. & Wang, K. (1993b) Nebulin as an actin ruler, J.Biol.Chem. 268, 20327-20334.

Kabsch, W., Mannherz, H. G., Suck, D., Pai, E. F. & Holmes, K. C. (1990) Atomic structure of the actin-DNase I complex, Nature. 347, 37-44.

Kruger, M., Wright, J. & Wang, K. (1991) Nebulin as a length regulator of thin filament length, nebulin size, and epitope profile., J.Cell Biol. 115, 97-107.

Labiet, S., Gibson, T., Lakey, A., Leonard, K., Zeviani, M., Knight, P., Wardale, J. & Trinick, J. (1991) Evidence that nebulin is a protein-ruler in muscle thin filaments., FEBS letters. 282, 313-316.

Luo, G., Zhang, J. Q., Nguyen, T. P., Herrera, A. H., Paterson, B. & Horowits, R. (1997) Complete cDNA sequence and tissue localization of N-RAP, a novel nebulin-related protein of striated muscle., Cell Mot.Cytoskel. 38, 75-90.

McElhinny, A. S., Kolmerer, B., Fowler, V. M., Labeit, S. & Gregorio, C. C. (2001) The N-terminal end of nebulin interacts with tropomodulin at the pointed ends of the thin filaments., J.Biol.Chem. 276, 583-592.

Meng, Y., Yasunaga, T. & Wakkabayashi, T. (1995) Determination of the protein components of native thin filaments isolated from natural actomyosin: Nebulin and a-actinin are associated with actin filaments., J.Biochem. 118, 422-427.

Moncman, C. L. & Wang, K. (1999) Functional dissection of nebulette demonstrates actin binding of nebulin-like repeats and Z-line targeting of SH3 and linker domains., Cell Motility Cytoskeleton. 44, 1-22.

Nwe, T. M., Maruyama, K. & Shimada, Y. (1999) Relation of nebulin and connectin (titin) to dynamics of actin in nascent myofibrils of cultured skeletal muscle cells., Exp.Cell Res. 252, 33-40.

Pelin, K., Hilpela, P., Donner, K., Sewry, C., Akkari, P. A., Wilton, S. D., Wattanasirichaigoon, D., Bang, M.-L., Centner, T., Henefeld, F., Odent, S., Fardeau, M., Urtizberea, J. A., Muntoni, F., Dubowitz, V., Beggs, A. H., Laing, N. G., Labeit, S., De La Chapelle, A. & Wallgren-Pettersson, C. (1999) Mutations in the nebulin gene associated with autosomal recessive nemaline myopathy., PNAS. 96, 2305-2310.

Pfuhl, M., Winder, S. J., Castiglione Morelli, M. A., Labeit, S. & Pastore, A. (1996) Correlation between conformational and binding properties of nebulin repeats, J.Mol.Biol. 257, 367-384.

Pfuhl, M., Winder, S. J. & Pastore, A. (1994) Nebulin, a helical actin binding protein, EMBO J. 13, 1782-1789.

Root, D. D. & Wang, K. (1994) Calmodulin-sensitive interaction of human nebulin fragments with actin and myosin, Biochemistry. 33, 12581-12591.

Tatsumi, R., Hattori, A. & Takahashi, K. (1992) Purification and characterization of nebulin subfragments produced by 0.1mM CaCl2., J.Biochem. 112, 780-785.

Tatsumi, R. & Takahashi, K. (1992) Calcium-induced fragmentation of skeletal muscle nebulin filaments, J.Biochem. 112, 775-777.

Wang, K., Kniper, M., Huang, Q.Q., van Heerden, A., Hsu, L., Gutierrez, G. (1996) J.Biol.Chem. 271, 4304-4314.

Wang, K. (1999) Nebulin. Guidebook to the Cytoskeleton Sambrook & Tooze, Oxford Uni.Press. p109-p111.

Wright, J., Huang, Q.-Q. & Wang, K. (1993) Nebulin is a full-length template of actin filaments in the skeletal muscle sarcomere: an immunoelectron microscope study of its orientation and span with site-directed monoclonal antibodies., J.Muscle Res.Cell Motility. 14, 476-483.

Zhang, J. Q., Weisenberg, A. & Horowits, R. (1998) Expression and purification of large nebulin fragments and their interaction with actin, Biophys.J. 74, 349-359.

Zhukarev, V., Sanger, J. M., Sanger, J. W., Goldman, Y. E. & Shuman, H. (1997) Distribution and orientation of rhodamine-phalloidin bound to thin filaments in skeletal and cardiac myofibrils, Cell Mot.Cytoskel. 37, 363-377.

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