Ca2+-induced tropomyosin movement in Limulus thin filaments revealed by three-dimensional reconstruction

Lehman, William; Craig, Roger; Vibert, Peter

doi:10.1038/368065a0

Cited by 317 publications

(333 citation statements)

References 23 publications

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“…Early work on which filament system was regulatory was contradictory, with some indicating that Limulus was only thin filament regulated and other that Limulus (and tarantula) muscle was dually regulated (Lehman and Szent-Györgyi, 1975;Chantler and Szent-Györgyi, 1978;Sellers et al, 1980). That there is a tropomyosin-troponin based thin filament regulatory system is unambiguous Lehman et al, 1976Lehman et al, , 1994Reedy et al, 1994b), although full movement of the tropomyosin to expose the entire myosin binding site on the thin filament requires both Ca ++ and myosin head binding to the thin filament . Limulus troponin C with rabbit troponin I and T restores full Ca ++ sensitivity to rabbit actomyosin (Lehman, 1975).…”

Section: Ecdysozoa Nematoda-mentioning

confidence: 99%

Invertebrate muscles: Thin and thick filament structure; molecular basis of contraction and its regulation, catch and asynchronous muscle

Hooper

Hobbs

Thuma

2008

Progress in Neurobiology

128

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This is the second in a series of canonical reviews on invertebrate muscle. We cover here thin and thick filament structure, the molecular basis of force generation and its regulation, and two special properties of some invertebrate muscle, catch and asynchronous muscle. Invertebrate thin filaments resemble vertebrate thin filaments, although helix structure and tropomyosin arrangement show small differences. Invertebrate thick filaments, alternatively, are very different from vertebrate striated thick filaments and show great variation within invertebrates. Part of this diversity stems from variation in paramyosin content, which is greatly increased in very large diameter invertebrate thick filaments. Other of it arises from relatively small changes in filament backbone structure, which results in filaments with grossly similar myosin head placements (rotating crowns of heads every 14.5 nm) but large changes in detail (distances between heads in azimuthal registration varying from three to thousands of crowns). The lever arm basis of force generation is common to both vetebrates and invertebrates, and in some invertebrates this process is understood on the near atomic level. Invertebrate actomyosin is both thin (tropomyosin:troponin) and thick (primarily via direct Ca ++ binding to myosin) filament regulated, and most invertebrate muscles are dually regulated. These mechanisms are well understood on the molecular level, but the behavioral utility of dual regulation is less so. The phosphorylation state of the thick filament associated giant protein, twitchin, has been recently shown to be the molecular basis of catch. The molecular basis of the stretch activation underlying asynchronous muscle activity, however, remains unresolved.

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Section: Ecdysozoa Nematoda-mentioning

confidence: 99%

Invertebrate muscles: Thin and thick filament structure; molecular basis of contraction and its regulation, catch and asynchronous muscle

Hooper

Hobbs

Thuma

2008

Progress in Neurobiology

128

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“…It is unclear why a modest increase in apparent affinity occurred instead, but the effect is small in any case. DISCUSSION The thin filament has at least three conformations: an inhibited state in the presence of EGTA, a Ca 2ϩ -induced state, and an active state observed in the presence of strongly binding myosin cross-bridges (35)(36)(37). These structures have been compared with three-dimensional reconstructions of myosin S1-decorated thin filaments (38,39), leading to the conclusion that tropomyosin interferes with the binding site for myosin S1 in the inhibited state and (to a lesser extent) in the Ca 2ϩ state but not in the active state.…”

Section: Mgatpase Activation As a Function Of The Free Ca 2ϩ Concentrmentioning

confidence: 99%

Cooperative Effect of Calcium Binding to Adjacent Troponin Molecules on the Thin Filament-Myosin Subfragment 1 MgATPase Rate

Butters

Tobacman

1997

Journal of Biological Chemistry

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The myosin subfragment 1 (S1) MgATPase rate was measured using thin filaments with known extents of Ca 2؉ binding controlled by varying the ratio of native cardiac troponin versus an inhibitory troponin with a mutation in the sole regulatory Ca 2؉ binding site of troponin C. Fractional MgATPase activation was less than the fraction of troponins that bound Ca 2؉ , implying a cooperative effect of bound Ca 2؉ on cross-bridge cycling. Addition of phalloidin did not alter cooperative effects between bound Ca 2؉ molecules in the presence or absence of myosin S1. When the myosin S1 concentration was raised sufficiently to introduce cooperative myosin-myosin effects, lower Ca 2؉ concentrations were needed to activate the MgATPase rate. MgATPase activation remained less than Ca 2؉ binding, implying a true, not just an apparent, increase in Ca 2؉ affinity. MgATPase activation by Ca 2؉ was more cooperative than could be explained by cooperativeness of overall Ca 2؉ binding, the discrepancy between Ca 2؉ binding and MgATPase activation, or interactions between myosins. The results suggest the thin filament-myosin S1 MgATPase cycle requires calcium binding to adjacent troponin molecules and that this binding is cooperatively promoted by a single cycling cross-bridge. This mechanism is a potential explanation for Ca 2؉ -mediated regulation of crossbridge kinetics in muscle fibers.Just as isometric tension is cooperatively activated by Ca 2ϩ , so is the cardiac thin filament-myosin S1 1 MgATPase rate, even under conditions where there is no cooperativity in myosin S1 binding (1, 2). A possible explanation for this behavior is that ATPase activation is proportional to Ca 2ϩ binding to the many TnCs on each thin filament and that this calcium binding is cooperative (3). We tested the idea that Ca 2ϩ binding and MgATPase activation are proportional and found to the contrary that they are not. Instead, fractional MgATPase activation was considerably less than fractional Ca 2ϩ binding and more closely paralleled the number of pairs of adjacent troponins with Ca 2ϩ bound to both.To accomplish the above experiment, we employ a constitutively inhibitory form of cardiac troponin containing an inactivating mutation of the sole regulatory site of TnC, site II (4). This troponin, designated CBMII-Tn, results in a low thin filament-myosin S1 MgATPase rate that is not increased by the addition of Ca 2ϩ , analogous to previous results in which a similar TnC mutant was exchanged into myofibrils or muscle fibers (5-7). CBMII-Tn binds to actin-tropomyosin with an affinity identical to that of normal troponin in the absence of Ca 2ϩ . This binding, which is very tight for both normal troponin and for 8,9), permits the present report in which thin filaments are assembled with defined mixtures of normal troponin and CBMII-Tn. In the presence of saturating Ca 2ϩ concentrations, such thin filaments exhibit a fractional saturation of the TnC regulatory sites that is experimentally controllable by varying the relative concentrations of the two forms ...

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“…Upon Ca 2+ binding to the single Ca 2+ binding site at the N-domain of cTnC, the regulatory processes occurring on the thin filament can be characterized by a series of structural changes in the thin filament proteins. These changes include an opening of the N-domain of cTnC (4,5), changes in the conformations of the inhibitory region (Ir) and the regulatory region (Rr) of cTnI (6,7), a switch of the Ir/Rr of cTnI from interacting with actin to interacting with cTnC (8), and movement of Tm on the actin surface (9). These transitions are the structural basis of thin filament regulation and result in force generation via strong interactions between actin and myosin head (S1).…”

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confidence: 99%

Effects of PKA Phosphorylation of Cardiac Troponin I and Strong Crossbridge on Conformational Transitions of the N-Domain of Cardiac Troponin C in Regulated Thin Filaments

Dong

Jayasundar

et al. 2007

Biochemistry

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Regulation of cardiac muscle function is initiated by binding of Ca 2+ to troponin C (cTnC) which induces a series of structural changes in cTnC and other thin filament proteins. These structural changes are further modulated by crossbridge formation and fine tuned by phosphorylation of cTnI. The objective of the present study is to use a new Förster Resonance Energy Transfer-based structural marker to distinguish structural and kinetic effects of Ca 2+ binding, crossbridge interaction and protein kinase A phosphorylation of cTnI on the conformational changes of the cTnC N-domain. The FRET-based structural marker was generated by attaching AEDANS to one cysteine of a doublecysteine mutant cTnC(13C/51C) as a FRET donor and attaching DDPM to the other cysteine as the acceptor. The doubly labeled cTnC mutant was reconstituted into the thin filament by adding cTnI, cTnT, tropomyosin and actin. Changes in the distance between Cys13 and Cys51 induced by Ca 2+ binding/dissociation were determined by FRET-sensed Ca 2+ titration and stopped-flow studies, and time-resolved fluorescence measurements. The results showed that the presence of both Ca 2+ and strong binding of myosin head to actin was required to achieve a fully open structure of the cTnC N-domain in regulated thin filaments. Equilibrium and stopped-flow studies suggested that strongly bound myosin head significantly increased the Ca 2+ sensitivity and changed the kinetics of the structural transition of the cTnC N-domain. PKA phosphorylation of cTnI impacted the Ca 2+ sensitivity and kinetics of the structural transition of the cTnC N-domain but showed no global structural effect on cTnC opening. These results provide an insight into the modulation mechanism of strong crossbridge and cTnI phosphorylation in cardiac thin filament activation/relaxation processes. Keywordsphosphorylation; cardiac troponin C; thin filament; FRET; Ca 2+ activation; kinetics Force development during cardiac muscle contraction is dependent upon the strong interactions between myosin and the actin filament. These interactions are governed by the regulatory

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Ca2+-induced tropomyosin movement in Limulus thin filaments revealed by three-dimensional reconstruction

Cited by 317 publications

References 23 publications

Invertebrate muscles: Thin and thick filament structure; molecular basis of contraction and its regulation, catch and asynchronous muscle

Invertebrate muscles: Thin and thick filament structure; molecular basis of contraction and its regulation, catch and asynchronous muscle

Cooperative Effect of Calcium Binding to Adjacent Troponin Molecules on the Thin Filament-Myosin Subfragment 1 MgATPase Rate

Effects of PKA Phosphorylation of Cardiac Troponin I and Strong Crossbridge on Conformational Transitions of the N-Domain of Cardiac Troponin C in Regulated Thin Filaments

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