Ala Scanning of the Inhibitory Region of Cardiac Troponin I

In skeletal and cardiac muscles, troponin (Tn), which resides on the thin filament, senses a change in intracellular Ca2+ concentration. Tn is composed of TnC, TnI, and TnT. Ca2+ binding to the regulatory domain of TnC removes the inhibitory effect by TnI on the contraction. The inhibitory region of cardiac TnI spans from residue 138 to 149. Upon Ca2+ activation, the inhibitory region is believed to be released from actin, thus triggering actin-activation of myosin ATPase. In this study, we created a series of Ala-substitution mutants of cTnI to delineate the functional contribution of each amino acid in the inhibitory region to myofilament regulation. We found that most of the point mutations in the inhibitory region reduced the ATPase activity in the presence of Ca2+, which suggests the same region also acts as an activator of the ATPase. The thin filaments can also be activated by strong myosin head (S1)-actin interactions. The binding of N-ethylmaleimide-treated myosin subfragment 1 (NEM-S1) to actin filaments mimics such strong interactions. Interestingly, in the absence of Ca2+ NEM-S1-induced activation of S1 ATPase was significantly less with the thin filaments containing TnI(T144A) than that with the wild-type TnI. However, in the presence of Ca2+, there was little difference in the activation of ATPase activity between these preparations. In skeletal and cardiac muscles, troponin (Tn), which resides on the thin filament, senses a change in intracellular Ca2+ concentration. Tn is composed of TnC, TnI, and TnT. Ca2+ binding to the regulatory domain of TnC removes the inhibitory effect by TnI on the contraction. The inhibitory region of cardiac TnI spans from residue 138 to 149. Upon Ca2+ activation, the inhibitory region is believed to be released from actin, thus triggering actin-activation of myosin ATPase. In this study, we created a series of Ala-substitution mutants of cTnI to delineate the functional contribution of each amino acid in the inhibitory region to myofilament regulation. We found that most of the point mutations in the inhibitory region reduced the ATPase activity in the presence of Ca2+, which suggests the same region also acts as an activator of the ATPase. The thin filaments can also be activated by strong myosin head (S1)-actin interactions. The binding of N-ethylmaleimide-treated myosin subfragment 1 (NEM-S1) to actin filaments mimics such strong interactions. Interestingly, in the absence of Ca2+ NEM-S1-induced activation of S1 ATPase was significantly less with the thin filaments containing TnI(T144A) than that with the wild-type TnI. However, in the presence of Ca2+, there was little difference in the activation of ATPase activity between these preparations. Striated muscle thin filaments exist in equilibrium among multiple states. Ca2+ binding to the regulatory domain of troponin C (TnC) 2The abbreviations used are: TnCtroponin CcTncardiac troponinfsTnfast skeletal troponinMOPS4-morpholinepropanesulfonic acidIAANS2-(4′-(iodoacetamido)anilino)-naphthalene-6-sulfonic acidNEMN-ethylmaleimideANOVAanalysis of variancewtwild typeDTTdithiothreitol. along the thin filaments and strong cross-bridge interactions with thick filaments are thought to shift the equilibrium. Ca2+ binds to the regulatory domain of TnC, which regulates the interaction of troponin I (TnI) with actin-tropomyosin (Tm) and TnC (1Gordon A.M. Homsher E. Regnier M. Physiol. Rev. 2000; 80: 853-924Crossref PubMed Scopus (1329) Google Scholar, 2Kobayashi T. Solaro R.J. Annu. Rev. Physiol. 2005; 67: 39-67Crossref PubMed Scopus (275) Google Scholar, 3Tobacman L.S. Annu. Rev. Physiol. 1996; 58: 447-481Crossref PubMed Scopus (460) Google Scholar). In the thin filaments, the inhibitory region of TnI (residues 104–115 of rabbit fast skeletal TnI (fsTnI) or 138–149 of mouse cardiac TnI (cTnI)) undergoes a structural transition depending on the Ca2+ state of TnC (4Kobayashi T. Kobayashi M. Gryczynski Z. Lakowicz J.R. Collins J.H. Biochemistry. 2000; 39: 86-91Crossref PubMed Scopus (37) Google Scholar, 5Xing J. Chinnaraj M. Zhang Z. Cheung H.C. Dong W.J. Biochemistry. 2008; 47: 13383-13393Crossref PubMed Scopus (19) Google Scholar). In the absence of Ca2+ at the regulatory site(s) of TnC, the inhibitory region interacts with actin to prevent activation of myosin ATPase activity. When Ca2+ binds to the regulatory site(s) of TnC, the switch region of TnI, which is located at the C terminus of the inhibitory region, interacts with the newly exposed hydrophobic patch of the N-terminal regulatory domain of TnC (6Li M.X. Spyracopoulos L. Sykes B.D. Biochemistry. 1999; 38: 8289-8298Crossref PubMed Scopus (245) Google Scholar, 7McKay R.T. Pearlstone J.R. Corson D.C. Gagné S.M. Smillie L.B. Sykes B.D. Biochemistry. 1998; 37: 12419-12430Crossref PubMed Scopus (49) Google Scholar, 8McKay R.T. Tripet B.P. Pearlstone J.R. Smillie L.B. Sykes B.D. Biochemistry. 1999; 38: 5478-5489Crossref PubMed Scopus (45) Google Scholar). This interaction causes the removal of the inhibitory region and the second actin-Tm binding region of TnI from the actin surface and allows actin to interact with myosin. In the presence of Ca2+ at the regulatory sites of TnC, the inhibitory region and the central helical region of TnC are mutually stabilized, according to the recent x-ray crystal structure of the core domain of the fsTn complex (9Vinogradova M.V. Stone D.B. Malanina G.G. Karatzaferi C. Cooke R. Mendelson R.A. Fletterick R.J. Proc. Natl. Acad. Sci. U.S.A. 2005; 102: 5038-5043Crossref PubMed Scopus (255) Google Scholar). The sequence variations in the N-terminal and the C-terminal regions of TnT, another component of the Tn complex, are known to alter the Ca2+ sensitivity of myofilament activity (10Pan B.S. Potter J.D. J. Biol. Chem. 1992; 267: 23052-23056Abstract Full Text PDF PubMed Google Scholar, 11Gomes A.V. Venkatraman G. Davis J.P. Tikunova S.B. Engel P. Solaro R.J. Potter J.D. J. Biol. Chem. 2004; 279: 49579-49587Abstract Full Text Full Text PDF PubMed Scopus (69) Google Scholar). In addition, TnT is involved in the Ca2+-dependent interaction of the Tn complex with actin-Tm (12Malnic B. Farah C.S. Reinach F.C. J. Biol. Chem. 1998; 273: 10594-10601Abstract Full Text Full Text PDF PubMed Scopus (94) Google Scholar). However, the molecular mechanism whereby TnT participates in the Ca2+ regulation has not been established. troponin C cardiac troponin fast skeletal troponin 4-morpholinepropanesulfonic acid 2-(4′-(iodoacetamido)anilino)-naphthalene-6-sulfonic acid N-ethylmaleimide analysis of variance wild type dithiothreitol. There is evidence supporting the idea that each amino acid residue in the inhibitory region of TnI contributes differently and to a different degree to myofilament activities. One example is genetic mutations and phosphorylation of amino acid residues in the inhibitory region of cardiac TnI that cause the modification of myofilament activities. In hypertrophic or restrictive cardiomyopathy-linked mutations found in the inhibitory region, such as R142Q, L145Q, and R146G/Q/W mutations (mouse cTnI sequence number), induce Ca2+ sensitization of myofilament activities and an increase in ATPase/tension at low [Ca2+] (13Chang A.N. Parvatiyar M.S. Potter J.D. Biochem. Biophys. Res. Commun. 2008; 369: 74-81Crossref PubMed Scopus (34) Google Scholar, 14Harada K. Morimoto S. Jpn. J. Physiol. 2004; 54: 307-318Crossref PubMed Scopus (28) Google Scholar). Recently we reported that thin filaments reconstituted with R146G or R146W mutant cTnI bind Ca2+ tighter than those with cTnI(wt) (15Kobayashi T. Solaro R.J. J. Biol. Chem. 2006; 281: 13471-13477Abstract Full Text Full Text PDF PubMed Scopus (99) Google Scholar). The Ca2+ sensitization may occur as a result of the destabilization of the off-state of the thin filaments due to the mutation introduced into the actin-Tm-interacting residue, i.e. Arg-146, of cTnI. On the other hand, is by C the of the phosphorylation of has not been of was to cause Ca2+ in in Kobayashi T. M. Homsher E. Solaro R.J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google there is a that phosphorylation of to Ca2+ S. J. J. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar). reported that of cTnI is activation of cardiac muscle K. Res. PubMed Scopus Google Scholar). in each a change in a amino acid in the inhibitory region of TnI different and on myofilament activities. of this is to the functional of the amino acid residues in the inhibitory region to the regulatory the functional of the amino acid residues we used PubMed Scopus Google Scholar, J. Biol. 1998; PubMed Scopus Google Scholar). the interactions by not alter the is to or is of the most amino and is found in and exposed We found that the inhibitory region of cTnI we is the and also that the C-terminal of the inhibitory region the state of the thin We also found that is involved in activation of ATPase activity in the absence of wild-type and mutant and in was with 1 and the and the was and and to of 1 and by the of to the was to a with 1 1 and with the same the was with the same that was used of 1 was with and The was 1 and 1 was to the to and was on a fast with 1 and was with a of mouse and as T. Solaro R.J. J. 2005; 38: Full Text Full Text PDF PubMed Scopus Google Scholar). mouse with a at the terminus was and with a of and as Kobayashi T. Solaro R.J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). we that the at the terminus of has on myofilament activity PubMed Scopus Google Scholar). was from with 1 1 1 and by point The with 1 and was The was and was on a fast was from rabbit skeletal muscle as by and S. J. Biol. Chem. Full Text PDF PubMed Google Scholar). was in the presence of was by of rabbit muscle myosin and on a as J. Biol. Chem. 1992; 267: Full Text PDF PubMed Google Scholar). residues of mutant and with of in the presence of 1 and The was by the of and the was by and a The was (15Kobayashi T. Solaro R.J. J. Biol. Chem. 2006; 281: 13471-13477Abstract Full Text Full Text PDF PubMed Scopus (99) Google Scholar). of Tn in a containing 1 1 and The was the same The was reduced to and to the was by and the to a with and The Tn complex was with a of to in the same thin filaments, we actin and by the Ca2+ binding we actin and in a by the of Tn to with of actin to was to Tn in the thin preparations. Tn an of change binding Ca2+ J.P. C. Kobayashi T. Solaro R.J. Tikunova S.B. Biophys. J. Full Text Full Text PDF PubMed Scopus Google the presence of Tn may with the of Ca2+ binding to thin that a of Tn and of Tn the same we also Ca2+ binding an of Tn with actin and a with with a containing a and a The Ca2+ binding was by of at of or of the was used to that the with a of Tn in the thin filaments the same as that of thin filaments with of The change was to be due to Ca2+ binding to the regulatory of in the The 1 and The was at and the Ca2+ was the C. S. 2004; PubMed Scopus Google Scholar). The as (15Kobayashi T. Solaro R.J. J. Biol. Chem. 2006; 281: 13471-13477Abstract Full Text Full Text PDF PubMed Scopus (99) Google Scholar, J.P. C. Kobayashi T. Solaro R.J. Tikunova S.B. Biophys. J. Full Text Full Text PDF PubMed Scopus Google Scholar). a of Ca2+ as from the The and are as a from The of Tn with Ca2+ binding and actin-Tm interaction was as in and are the Ca2+ the Tn complex and the thin filaments, was by is a of and and and are with and The actin, myosin and Tn in and or at was by the of a of 1 ATPase activity was from the of a T. K. K. J. Biochem. PubMed Scopus Google Scholar). ATPase activity was from to In the of ATPase actin at as E. Proc. Natl. Acad. Sci. U.S.A. PubMed Scopus Google Scholar, Kobayashi T. Biophys. J. 2008; Full Text Full Text PDF PubMed Scopus Google and ATPase S1 concentration. In and the S1 has been from the and the is as a from or was by by the as a multiple was that the is of the most and and Google Scholar). residues from to was to to the of at each be that the of each of these residues to the structure of the inhibitory region the structure of the switch region, which the inhibitory region, the structural by the L. Biol. PubMed Scopus Google Scholar). We Ca2+ binding to the Tn complex with mutation of cTnI in the inhibitory region residues or in reconstituted thin The Ca2+ binding was reported by change of to of the we reported to of the mutant Ca2+ binding as as Ca2+ from the regulatory of and of the complex to those with wild-type and J.P. C. Kobayashi T. Solaro R.J. Tikunova S.B. Biophys. J. Full Text Full Text PDF PubMed Scopus Google Scholar). at of Ca2+ binding to the reconstituted thin filaments with the same as from (15Kobayashi T. Solaro R.J. J. Biol. Chem. 2006; 281: 13471-13477Abstract Full Text Full Text PDF PubMed Scopus (99) Google Scholar, B.S. Solaro R.J. J. Biol. Chem. Full Text PDF PubMed Google Scholar, L.S. J. Biol. Chem. Full Text PDF PubMed Google Scholar). this can be used not the Ca2+ binding the complex also the thin filaments as This to from different regulatory Ca2+ binding to the Tn complex a of the Ca2+ binding to the thin filaments the This that the of to are different in the Tn complex and the thin filaments in the presence of In the thin filaments, may the actin-Tm surface that a less exposed Ca2+ the reconstituted thin filaments, we also the Ca2+ binding with to as and The Ca2+ binding of the Tn and the reconstituted thin filaments are in The in the of the regulatory with mutant cTnI from that with wild type from the and are in Ca2+ binding used to the of the interaction of the inhibitory region with other thin by at each in the residues from to the effect of mutations of cTnI on the Ca2+ binding the different regulatory from cTnI(wt) by by a from cTnI(wt) by by a from cTnI(wt) by by a from cTnI(wt) by by a from cTnI(wt) by by a from cTnI(wt) by by a from cTnI(wt) by by a from cTnI(wt) by by a not from cTnI(wt) by by a from cTnI(wt) by by a from cTnI(wt) by by a not in a of mutation on from from Ca2+ binding the Tn and the thin filaments as in from cTnI(wt) by by The of of of these residues on Ca2+ binding to the Tn complex are in 1 and and mutations not Ca2+ binding of the Tn with the residues from of of the amino acid residues in the in Ca2+ of the Tn The mutation the each of the of the mutations there of Ca2+ binding of the Tn complex by of amino acid residues in the inhibitory In on Ca2+ binding of the thin filaments 1 and than those of the that Ca2+ binding to the thin filaments in sensitization of the to Ca2+, as by of in The mutation significantly the Ca2+ binding of the thin filaments by and as as also sensitization of the thin filaments to Ca2+, these not significantly different from cTnI(wt) with analysis cTnI(wt) with the The mutation not alter the Ca2+ binding of the thin The thin filaments with the mutation not a change with we not Ca2+ binding to the thin filaments with Ca2+ binding to the Tn complex and actin-Tm binding are Ca2+ binding to the Tn complex The Ca2+ binding the Tn and the thin filaments used to between Ca2+ and actin-Tm each Tn complex The is a of of the interaction of the Tn complex with actin-Tm and was as and be that the thin filaments exist in equilibrium between at which different Ca2+ binding is of the thin filaments the be Ca2+-dependent interactions of the Tn complex with actin-Tm can also be a However, may as J.P. L.S. J. Biol. Chem. 1992; 267: Full Text PDF PubMed Google Scholar). from the Ca2+-dependent interaction of the Tn complex with actin-Tm was by mutations in the of the inhibitory region and The mutation a in the Ca2+-dependent we not the Ca2+ binding of the thin filaments with the we not the the of myosin activities is in The of the wild-type Tn to the complex of myosin actin, and the ATPase activity to from in the presence of Ca2+ the activity to in the absence of Ca2+ the ATPase activity was by Ca2+ in the presence of the Tn In the presence of Ca2+, the mutation in this in a of ATPase activity with cTnI(wt) the of the inhibitory region in Ca2+ and mutations Ca2+ ATPase activities with of these mutations is that the mutation Ca2+ activation the Ca2+ binding In the shift of the equilibrium of the thin to be by the of the Ca2+ binding this the mutation In the absence of Ca2+, most of the mutations the inhibitory The mutation the effect cTnI in this study, the ATPase activity to in Ca2+ and mutations effect on the ATPase activity in the absence of The thin filaments can be activated a strong cross-bridge We used to such an S1 has little ATPase activity by binds to actin in the presence of and the state of the thin of to the ATPase the thin filaments and ATPase activity. the of activation of ATPase with mutations those with the ATPase In the of the ATPase to with 1 in the presence of Ca2+ in and to in the absence of Ca2+ in of ATPase be in the presence and absence of reported the mutation not Ca2+-dependent activity The of activity with mutation as as that with wild-type Tn in the presence of in the absence of Ca2+, ATPase activity with was not activated as as ATPase with wild-type TnI by to 1 ATPase activities ATPase activity was from that with wild-type TnI in the absence of Ca2+ not of the of the thin filaments with In the of which introduced a reduced ATPase activity in the presence of Ca2+ and an activity in the absence of Ca2+ with wild type an of ATPase activity to the same as that with wild type in the presence of Ca2+ On the other hand, in the absence of Ca2+, ATPase activities with or the same as those with wild these the of the state of the thin filaments by the In the of the mutations of the of the inhibitory region to ATPase the same as those with cTnI(wt) with in the absence of Ca2+ In the presence of Ca2+ and ATPase than those with This the destabilization of state of the thin filaments by these mutations and is also with the the in reported are the to the of the of the inhibitory region of cTnI on the Ca2+-dependent and the strong myofilament activities reconstituted We the of the inhibitory region is the Ca2+-dependent interaction of the Tn complex with the that we is the cTnI inhibitory the is Ca2+ activation, and most is involved in strong activation of the thin filaments in the presence of mutation not inhibitory in the absence of Ca2+ activation in the presence of Ca2+ The inhibitory region was to interact with actin in the absence of Ca2+ and the strong interaction between actin and myosin. One molecular mechanism the inhibitory was by J. Biol. Chem. 2005; Full Text Full Text PDF PubMed Scopus Google Scholar). on the the actin surface between a from the inhibitory region of TnI and the from that the inhibitory region interacts with actin that the actin undergoes a change that the actin in a to a surface less complex with myosin. On the other hand, of the thin filaments from suggests of myosin binding to actin by with the interaction between the inhibitory region and actin the of actin to prevent actin from interaction with myosin R. J. Biol. PubMed Scopus Google Scholar, M.V. Fletterick R.J. R. L.S. C. J. Biol. 2006; PubMed Scopus Google Scholar, P. R. J. Biol. PubMed Scopus Google Scholar). of the molecular mechanism the inhibitory by the inhibitory region of TnI, the equilibrium between the state of the thin filaments and the state of the thin filaments to be R. B. K. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus (37) Google Scholar, C. R. L.S. J. Biol. 2005; PubMed Scopus (99) Google Scholar). We found that of Ca2+-dependent interaction of Tn to actin-Tm by of amino acid residues in the inhibitory region are to However, ATPase activities by that the with the Ca2+ binding of the Tn complex and those with the thin the on the state and state of the thin filaments, the in state are the of the Ca2+-dependent interaction This is with other that hypertrophic cardiomyopathy-linked mutations found in the inhibitory region not the of the Tn complex to actin-Tm myofilament activity in the absence of Ca2+ is K. C.S. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar, R. A.V. J. T. Potter J.D. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, Kobayashi T. Biophys. J. Full Text Full Text PDF PubMed Scopus Google Scholar). this effect of the inhibitory region is in to the with the mutation in the second actin-Tm binding The second actin-Tm binding of TnI is located in the C-terminal domain of TnI. The C-terminal domain to a structure binds to actin-Tm in the absence of Ca2+ in and cTnI K. T. M. T. J. Biol. 2005; PubMed Scopus Google Scholar, Engel P. C. R. L.S. J. Biol. 2008; PubMed Scopus Google Scholar). K. T. M. T. J. Biol. 2005; PubMed Scopus Google the structure of the C-terminal domain of and the structure of the C-terminal domain to of the thin In structural the second actin-Tm binding spans residues of and of mouse cTnI. The from the C terminus of as most of the mutations of the inhibitory region in this study, in a sensitization to Ca2+ of the not the of ATPase activity B. S. R. R.J. C. K. D.B. A.M. C. C. J. Res. 2006; PubMed Scopus Google Scholar, K. Kobayashi T. Solaro R.J. J. Biol. Chem. 2008; Full Text Full Text PDF PubMed Scopus Google Scholar). This is of the in in mutation from the inhibitory region, R146G (mouse a of Ca2+-dependent mutations from the C-terminal and Ca2+-dependent regulation K. C. S. C. Biochem. J. PubMed Scopus Google Scholar, J. B. A.M. T. Regnier M. Physiol. PubMed Scopus Google Scholar). the C-terminal domain of cTnI the same structure as that of by K. T. M. T. J. Biol. 2005; PubMed Scopus Google and of cTnI are to be involved in the actin these may the functional difference between the inhibitory region and the second actin-Tm binding of TnI. the of Ca2+-dependent interaction of Tn to actin-Tm in not the degree of of the inhibitory of ATPase activity in the absence of Ca2+ in the mutation that the Ca2+-dependent interaction of Tn with not the inhibitory activity. in also that the of the ATPase activity at low [Ca2+] the inhibitory region of TnI, the degree of the of mutations on the inhibitory from residue to and J. Biol. Chem. Full Text PDF PubMed Google the inhibitory region of with to the inhibitory region and the ATPase not TnT and TnC, also found a of the amino acid residues that the of ATPase activity. the of the interaction of the inhibitory region with actin at low [Ca2+] is the interaction with TnC at [Ca2+] has been and The C-terminal of the inhibitory region, than the inhibitory region was as the Ca2+-dependent switch region, which interacts with the regulatory domain of TnC in a Ca2+-dependent (6Li M.X. Spyracopoulos L. Sykes B.D. Biochemistry. 1999; 38: 8289-8298Crossref PubMed Scopus (245) Google Scholar, 7McKay R.T. Pearlstone J.R. Corson D.C. Gagné S.M. Smillie L.B. Sykes B.D. Biochemistry. 1998; 37: 12419-12430Crossref PubMed Scopus (49) Google Scholar, 8McKay R.T. Tripet B.P. Pearlstone J.R. Smillie L.B. Sykes B.D. Biochemistry. 1999; 38: 5478-5489Crossref PubMed Scopus (45) Google Scholar). In the of fast skeletal most of the Ca2+-dependent binding between TnC and TnI to from the interaction between the switch region of TnI and the N-terminal regulatory domain of the inhibitory region, on the other hand, the crystal structure of the core domain of fsTn complex in the presence of Ca2+ that interacts with the central helical region of TnC (9Vinogradova M.V. Stone D.B. Malanina G.G. Karatzaferi C. Cooke R. Mendelson R.A. Fletterick R.J. Proc. Natl. Acad. Sci. U.S.A. 2005; 102: 5038-5043Crossref PubMed Scopus (255) Google Scholar). cardiac the of TnC was and the inhibitory region was not in the crystal structure of the core domain of complex S. K. PubMed Scopus Google x-ray of the cardiac thin filaments an structure of the region of TnC K. K. S. J. Biol. 2004; PubMed Scopus Google Scholar). in the region of the Tn complex and a interaction between the inhibitory region of TnI and the central of However, in the inhibitory region S1 ATPase activity in the presence of a a of to the thin filaments, we that cardiomyopathy-linked mutations found in the inhibitory region of R146G and R146W (mouse sequence the functional state of the thin filaments S. K. PubMed Scopus Google Scholar). The between the Ca2+ and the ATPase found in this also an state with a activity. In the of the inhibitory region the state of the thin filaments to the structure of the central is involved in the activation as by and PubMed Scopus Google or mutation of the of the inhibitory region the structural of the N-terminal regulatory domain of which may be a of myofilament activity Cheung H.C. Dong Biophys. J. 2008; Full Text Full Text PDF PubMed Scopus Google to be In this study, we not the with the the thin filaments with the mutation not the change with However, as in the mutation the activity in the absence of Ca2+, this mutation the interaction between the inhibitory region of cTnI and actin in the absence of the mutation in a in the activity in the presence of we in Kobayashi T. Biophys. J. Full Text Full Text PDF PubMed Scopus Google a mutation that causes a and an increase of the ATPase activity in the presence and the absence of Ca2+, is to the functional state of the thin This is with on the activation of ATPase activity in which a of the state of the thin filaments as The thin filaments with a mutation that the state a Ca2+ (15Kobayashi T. Solaro R.J. J. Biol. Chem. 2006; 281: 13471-13477Abstract Full Text Full Text PDF PubMed Scopus (99) Google the Ca2+ binding of the each state of the thin filaments not been is that the mutation the Ca2+-dependent interaction with actin-Tm and the The mutation little on Ca2+-dependent interaction of Tn with actin-Tm and on activities in the presence and absence of Ca2+ However, the mutation activation and S1 binding of the thin filaments in the absence of Ca2+ and This is most due to a reduced of S1 the off-state of the thin filaments with modification of such as phosphorylation or with another amino result in an of Ca2+ sensitivity of myofilament strong cross-bridge interaction can these In of the inhibitory region of cTnI that is the and This suggests the presence of an state of the thin filaments that the activity. We also found that which effect on Ca2+-dependent ATPase activity to is involved in strong activation of ATPase in the absence of We myosin S1 and We also R. Solaro and of the