Myosin subfragment-1 is sufficient to move actin filaments in vitro

Nature ◽  
1987 ◽  
Vol 328 (6130) ◽  
pp. 536-539 ◽  
Author(s):  
Yoko Yano Toyoshima ◽  
Stephen J. Kron ◽  
Elizabeth M. McNally ◽  
Kenneth R. Niebling ◽  
Chikashi Toyoshima ◽  
...  
Keyword(s):  
Actin Filaments ◽  
Myosin Subfragment ◽  
Myosin Subfragment 1

scholarly journals Polymerization of actin. VI. The polarity of the actin filaments in the acrosomal process and how it might be determined.

1979 ◽  
Vol 81 (3) ◽  
pp. 608-623 ◽  
Author(s):  
L G Tilney ◽  
N Kallenbach
Keyword(s):  
Low Temperature ◽  
Actin Filaments ◽  
Critical Concentration ◽  
Actin Monomer ◽  
Spontaneous Nucleation ◽  
Myosin Subfragment ◽  
Acrosomal Reaction ◽  
Myosin Subfragment 1

The polarity of the actin filaments which assemble from the nucleating body or actomere of Thyone and Pisaster sperm was determined using myosin subfragment 1 decoration. The polarity was found to be unidirectional with the arrowheads pointing towards the cell center. When polymerization is induced at low temperature with concentrations of actin near the critical concentration for polymerization, elongation of filaments occurs preferentially off the apical end. If the sperm are induced to undergo the acrosomal reaction with an ionophore, the polarity of the actin filaments attached to the actomere is the same as that already described, but the filaments which polymerize parallel to, but peripheral to, those extending from the actomere are randomly polarized. These randomly polarized filaments appear to result from spontaneous nucleation. When sperm are induced to undergo the acrosomal reaction with eggs, the polarity of the actin filaments is also unidirectional with the arrowheads pointing towards the cell center. From these results we conclude: (a) that the actomere, by nucleating the polymerization of actin filaments, controls the polarity of the actin filaments in the acrosomal process, (b) that the actomere recognizes a surface of the actin monomer that is different from that surface recognized by the dense material attached to membranes, and (c) that egg myosin could not act to pull the sperm into the egg. Included is a discussion of how the observation that monomers add largely to one end of a decorated filament in vitro relates to these in vivo observations.

Structure of Myosin Subfragment-1 from Electron Microscopy and Crystallography

1988 ◽  
Vol 46 ◽  
pp. 156-157
Author(s):  
Donald A. Winkelmann
Keyword(s):  
Electron Microscopy ◽  
Molecular Weight ◽  
Actin Filaments ◽  
Light Chains ◽  
Myosin Filament ◽  
Primary Role ◽  
Heavy Chains ◽  
Myosin Subfragment ◽  
Myosin Subfragment 1 ◽  
Globular Head

The primary role of the interaction of actin and myosin is the generation of force and motion as a direct consequence of the cyclic interaction of myosin crossbridges with actin filaments. Myosin is composed of six polypeptides: two heavy chains of molecular weight 220,000 daltons and two pairs of light chains of molecular weight 17,000-23,000. The C-terminal portions of the myosin heavy chains associate to form an α-helical coiled-coil rod which is responsible for myosin filament formation. The N-terminal portion of each heavy chain associates with two different light chains to form a globular head that binds actin and hydrolyses ATP. Myosin can be fragmented by limited proteolysis into several structural and functional domains. It has recently been demonstrated using an in vitro movement assay that the globular head domain, subfragment-1, is sufficient to cause sliding movement of actin filaments.The discovery of conditions for crystallization of the myosin subfragment-1 (S1) has led to a systematic analysis of S1 structure by x-ray crystallography and electron microscopy. Image analysis of electron micrographs of thin sections of small S1 crystals has been used to determine the structure of S1 in the crystal lattice.

Myosin subfragment 1 binding to relaxed actin filaments and steric model of relaxation

Biochemistry ◽  
1981 ◽  
Vol 20 (3) ◽  
pp. 641-649 ◽  
Author(s):  
John M. Murray ◽  
Annemarie Weber ◽  
Mary K. Knox
Keyword(s):  
Actin Filaments ◽  
Myosin Subfragment ◽  
Myosin Subfragment 1

scholarly journals Regulation of the acto·myosin subfragment 1 interaction by troponin/tropomyosin. Evidence for control of a specific isomerization between two acto·myosin subfragment 1 states

1991 ◽  
Vol 279 (3) ◽  
pp. 711-718 ◽  
Author(s):  
D F A McKillop ◽  
M A Geeves
Keyword(s):  
Skeletal Muscle ◽  
Active Site ◽  
Actin Filaments ◽  
Thin Filaments ◽  
Closed State ◽  
Myosin Subfragment ◽  
Actomyosin Interaction ◽  
Binding Isotherms ◽  
Myosin Subfragment 1 ◽  
Site Binding

The co-operative binding of myosin subfragment 1 (S1) to reconstituted skeletal-muscle thin filaments has been examined by monitoring the fluorescence of a pyrene probe on Cys-374 of actin. The degree of co-operativity differs when phosphate, sulphate or ADP are bound to the S1 active site. Binding isotherms have been analysed according to the Geeves & Halsall [(1987) Biophys. J. 52, 215-220] model, which proposed that troponin and tropomyosin effected regulation of the actomyosin interaction by controlling an isomerization of the actomyosin complex. The data support the proposal that seven actin monomers associated with a single tropomyosin molecule act as a co-operative unit that can be in one of two states. In the ‘closed’ state myosin can bind to actin, but the subsequent isomerization is prevented. The isomerization is only allowed after the seven-actin unit is in the ‘open’ form. Ca2+ controls the proportion of actin filaments in the ‘closed’ and ‘open’ forms in the absence of myosin heads. The ratio of ‘closed’ to ‘open’ forms is approx. 50:1 in the absence of Ca2+ and 5:1 in its presence.

scholarly journals Molecular Mechanisms of Muscle Weakness Associated with E173A Mutation in Tpm3.12. Troponin Ca2+ Sensitivity Inhibitor W7 Can Reduce the Damaging Effect of This Mutation

2020 ◽  
Vol 21 (12) ◽  
pp. 4421
Author(s):  
Yurii S. Borovikov ◽  
Armen O. Simonyan ◽  
Stanislava V. Avrova ◽  
Vladimir V. Sirenko ◽  
Charles S. Redwood ◽  
...  
Keyword(s):  
Atpase Activity ◽  
Muscle Weakness ◽  
Conformational Changes ◽  
Molecular Mechanisms ◽  
Spatial Arrangement ◽  
Muscle Fibres ◽  
Thin Filaments ◽  
Myosin Subfragment ◽  
Myosin Subfragment 1

Substitution of Ala for Glu residue in position 173 of γ-tropomyosin (Tpm3.12) is associated with muscle weakness. Here we observe that this mutation increases myofilament Ca2+-sensitivity and inhibits in vitro actin-activated ATPase activity of myosin subfragment-1 at high Ca2+. In order to determine the critical conformational changes in myosin, actin and tropomyosin caused by the mutation, we used the technique of polarized fluorimetry. It was found that this mutation changes the spatial arrangement of actin monomers and myosin heads, and the position of the mutant tropomyosin on the thin filaments in muscle fibres at various mimicked stages of the ATPase cycle. At low Ca2+ the E173A mutant tropomyosin shifts towards the inner domains of actin at all stages of the cycle, and this is accompanied by an increase in the number of switched-on actin monomers and myosin heads strongly bound to F-actin even at relaxation. Contrarily, at high Ca2+ the amount of the strongly bound myosin heads slightly decreases. These changes in the balance of the strongly bound myosin heads in the ATPase cycle may underlie the occurrence of muscle weakness. W7, an inhibitor of troponin Ca2+-sensitivity, restores the increase in the number of myosin heads strongly bound to F-actin at high Ca2+ and stops their strong binding at relaxation, suggesting the possibility of using Ca2+-desensitizers to reduce the damaging effect of the E173A mutation on muscle fibre contractility.

scholarly journals The visualization of actin filament polarity in thin sections. Evidence for the uniform polarity of membrane-associated filaments.

1978 ◽  
Vol 79 (3) ◽  
pp. 846-852 ◽  
Author(s):  
D A Begg ◽  
R Rodewald ◽  
L I Rebhun
Keyword(s):  
Actin Filament ◽  
Actin Filaments ◽  
Intestinal Epithelial ◽  
Improved Method ◽  
Thin Sections ◽  
Myosin Subfragment ◽  
Brush Borders ◽  
Nonmuscle Cells ◽  
Myosin Subfragment 1 ◽  
Uniform Polarity

We have developed an improved method for visualizing actin filament polarity in thin sections. Myosin subfragment-1 (S-1)-decorated actin filaments display a dramatically enhanced arrowhead configuration when fixed in a medium which contains 0.2 % tannic acid. With the exception of brush borders from intestinal epithelial cells, the arrowhead periodicity of decorated filaments in a variety of nonmuscle cells is similar to that in isolated myofibrils. The periodicity of decorated filaments in brush borders is significantly smaller. Actin filaments which attach to membranes display a clear, uniform polarity, with the S-1 arrowheads pointing away from the plasma membrane, while those which comprise the stress fibers of myoblasts and CHO cells have antiparallel polarities. These observations are consistent with a sliding filament mechanism of cell motility.

scholarly journals A membrane cytoskeleton from Dictyostelium discoideum. III. Plasma membrane fragments bind predominantly to the sides of actin filaments.

1984 ◽  
Vol 99 (1) ◽  
pp. 71-78 ◽  
Author(s):  
C M Goodloe-Holland ◽  
E J Luna
Keyword(s):  
Plasma Membrane ◽  
Membrane Proteins ◽  
Dictyostelium Discoideum ◽  
Electron Micrographs ◽  
Actin Filaments ◽  
Lateral Movement ◽  
Heavy Meromyosin ◽  
Membrane Cytoskeleton ◽  
Myosin Subfragment ◽  
Myosin Subfragment 1

The binding between sonicated Dictyostelium discoideum plasma membrane fragments and F-actin on Sephacryl S-1000 beads was found to be competitively inhibited by myosin subfragment-1. This inhibition is MgATP-sensitive, exhibits a Ki of approximately 5 X 10(-8) M, and is reciprocal, since membranes inhibit the binding of 125I-heavy meromyosin to F-actin on beads. These experiments demonstrate that membrane binding and S-1 binding to F-actin on beads are mutually exclusive and, therefore, that the membrane fragments bind predominantly to the sides, rather than to the ends, of the actin filaments. This conclusion is supported by electron micrographs that show many lateral associations between membrane fragments and bead-associated actin filaments. Such lateral associations could play an important role in the organization and lateral movement of membrane proteins by the cytomusculature.

scholarly journals Synthetic thick filaments: A new avenue for better understanding the myosin super-relaxed state in healthy, disease, and mavacamten-treated cardiac systems

2020 ◽  
pp. jbc.RA120.016506 ◽  
Author(s):  
Sampath K Gollapudi ◽  
Ming Yu ◽  
Qing-Fen Gan ◽  
Suman Nag
Keyword(s):  
Clinical Stage ◽  
Thick Filament ◽  
Functional Evaluation ◽  
Thick Filaments ◽  
Myosin Subfragment ◽  
Molecule Inhibitor ◽  
Myosin Subfragment 1 ◽  
Vitro Model ◽  
Low Ionic Strength

A hallmark feature of myosin-II is that it can spontaneously self-assemble into bipolar synthetic thick filaments (STFs) in low ionic strength buffers, thereby serving as a reconstituted in-vitro model for muscle thick filament. While these STFs have been extensively used for structural characterization, their functional evaluation has been limited.  In this report, we show that myosins in STFs mirror the more electrostatic and cooperative interactions that underlie the energy-sparing super-relaxed (SRX) state, which are not seen using shorter myosin sub-fragments, heavy meromyosin (HMM) and myosin subfragment-1 (S1). Using these STFs, we show several pathophysiological insults in hypertrophic cardiomyopathy, including the R403Q myosin mutation, phosphorylation of myosin light chains, and increased ADP:ATP ratio destabilize the SRX population. Furthermore, wild-type myosin containing STFs, but not S1, HMM, or STFs-containing R403Q myosin, recapitulated the ADP-induced destabilization of the SRX state. Studies involving a clinical-stage small molecule inhibitor, mavacamten, showed that it is not only more effective in increasing myosin SRX population in STFs than in S1 or HMM ,  but it also increases myosin SRX population equally well in STFs made of healthy and disease-causing R403Q myosin. Importantly, we also found that pathophysiological perturbations   such as elevated ADP concentration weakens the mavacamten’s ability to increase the myosin SRX population, suggesting that mavacamten-bound myosin heads are not permanently protected in the SRX state but can be recruited   into action. These findings collectively emphasize that STFs serve as a valuable tool to provide novel insights into the myosin SRX state in healthy, disease, and therapeutic conditions.

Corrections - Myosin Subfragment 1 Binding to Relaxed Actin Filaments and Steric Model of Relaxation

Biochemistry ◽  
1981 ◽  
Vol 20 (17) ◽  
pp. 5094-5094
Author(s):  
John Murray ◽  
Annemarie Weber ◽  
Mary Knox
Keyword(s):  
Actin Filaments ◽  
Myosin Subfragment ◽  
Myosin Subfragment 1
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