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.
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 Ca(2+) concentrations, fn3 fragments significantly enhance active tension. This effect is most pronounced at short sarcomere length, and as a result the length-dependence of Ca(2+) activation is reduced. A model of how titin's fn3-like domains may influence actomyosin interaction is proposed.
Muhle-Goll
C
C
European Molecular Biology Laboratory, Meyerhofstr. 1, Heidelberg, D-69117, Germany. muhle@embl-heidelberg.de
Habeck
M
M
Cazorla
O
O
Nilges
M
M
Labeit
S
S
Granzier
H
H
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000494:pharmacology; D064211:Connectin; D017124:Conserved Sequence; D005353:Fibronectins / Q000737:chemistry*; D006352:Heart Ventricles / Q000166:cytology / Q000187:drug effects; D006801:Humans; D007136:Immunoglobulins / Q000737:chemistry; D051379:Mice; D008807:Mice, Inbred BALB C; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000737:chemistry* / Q000378:metabolism* / Q000494:pharmacology; D009200:Myocardial Contraction / Q000187:drug effects; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D015879:Myosin Subfragments / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D011485:Protein Binding; D011494:Protein Kinases / Q000737:chemistry* / Q000378:metabolism* / Q000494:pharmacology; D017434:Protein Structure, Tertiary; D011817:Rabbits; D012518:Sarcomeres / Q000187:drug effects / Q000378:metabolism; D016415:Sequence Alignment; D012997:Solvents / Q000378:metabolism; D013329:Structure-Activity Relationship; D016276:Ventricular Function
D064211:Connectin; D005353:Fibronectins; D007136:Immunoglobulins; D009124:Muscle Proteins; D015879:Myosin Subfragments; D010446:Peptide Fragments; D012997:Solvents; C577285:TTN protein, human; D011494:Protein Kinases; D002118:Calcium