TY - JOUR
T1 - Structural and Functional Studies of Titin's fn3 Modules Reveal Conserved Surface Patterns and Binding to Myosin S1 - A Possible Role in the Frank-Starling Mechanism of the Heart
AU - Muhle-Goll, Claudia
AU - Habeck, Michael
AU - Cazorla, Olivier
AU - Nilges, Michael
AU - Labeit, Siegfried
AU - Granzier, Henk
N1 - Funding Information:
Rabbit myosin and fragments, F-actin, troponin and tropomyosin were obtained as a gift from Dr B. Bullard. Drs P. A. Kenny and D. G. Higgins made their multiple sequence alignment of all muscle fn3 domains available to us. We are indebted to G. Stier for supplying pET vectors and TEV protease. The plasmid encoding Z1Z2 was obtained as a gift from T. Centner. We gratefully acknowledge the support by the Deutsche Forschungsgemeinschaft (MU 1606/1-1 to C.M.G.; NI 499/1-2 to M.N.; La668/5-2 to S.L.), and the NIH (HL61497 and HL62881 to H. G.).
PY - 2001/10/19
Y1 - 2001/10/19
N2 - The A-band part of titin, a striated-muscle specific protein spanning from the Z-line to the M-line, mainly consists of a well-ordered super-repeat array of immunoglobulin-like and fibronectin-type III (fn3)-like domains. Since it has been suspected that the fn3 domains might represent titin's binding sites to myosin, we have developed structural models for all of titin's 132 fn3-like domains. A subset of eight experimentally determined fn3 structures from a range of proteins, including titin itself, was used as homology templates. After grouping the models according to their position within the super-repeat segment of the central A-band titin region, we analyzed the models with respect to side-chain conservation. This showed that conserved residues form an extensive surface pattern predominantly at one side of the domains, whereas domains outside the central C-zone super-repeat region show generally less conserved surfaces. Since the conserved surface residues may function as protein-binding sites, we experimentally studied the binding properties of expressed multi-domain fn3 fragments. This revealed that fn3 fragments specifically bind to the sub-fragment 1 of myosin. We also measured the effect of fn3 fragments on the contractile properties of single cardiac myocytes. At sub-maximal Ca2+ concentrations, fn3 fragments significantly enhance active tension. This effect is most pronounced at short sarcomere length, and as a result the length-dependence of Ca2+ activation is reduced. A model of how titin's fn3-like domains may influence actomyosin interaction is proposed.
AB - The A-band part of titin, a striated-muscle specific protein spanning from the Z-line to the M-line, mainly consists of a well-ordered super-repeat array of immunoglobulin-like and fibronectin-type III (fn3)-like domains. Since it has been suspected that the fn3 domains might represent titin's binding sites to myosin, we have developed structural models for all of titin's 132 fn3-like domains. A subset of eight experimentally determined fn3 structures from a range of proteins, including titin itself, was used as homology templates. After grouping the models according to their position within the super-repeat segment of the central A-band titin region, we analyzed the models with respect to side-chain conservation. This showed that conserved residues form an extensive surface pattern predominantly at one side of the domains, whereas domains outside the central C-zone super-repeat region show generally less conserved surfaces. Since the conserved surface residues may function as protein-binding sites, we experimentally studied the binding properties of expressed multi-domain fn3 fragments. This revealed that fn3 fragments specifically bind to the sub-fragment 1 of myosin. We also measured the effect of fn3 fragments on the contractile properties of single cardiac myocytes. At sub-maximal Ca2+ concentrations, fn3 fragments significantly enhance active tension. This effect is most pronounced at short sarcomere length, and as a result the length-dependence of Ca2+ activation is reduced. A model of how titin's fn3-like domains may influence actomyosin interaction is proposed.
KW - Fibronectin type III
KW - Molecular modeling
KW - Muscle contraction
KW - Striated muscle
KW - Titin
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U2 - 10.1006/jmbi.2001.5017
DO - 10.1006/jmbi.2001.5017
M3 - Article
C2 - 11800567
AN - SCOPUS:0035914471
SN - 0022-2836
VL - 313
SP - 431
EP - 447
JO - Journal of Molecular Biology
JF - Journal of Molecular Biology
IS - 2
ER -