scholarly journals Structural changes induced in Ca2+-regulated myosin filaments by Ca2+ and ATP.

1989 ◽  
Vol 109 (2) ◽  
pp. 529-538 ◽  
Author(s):  
L L Frado ◽  
R Craig
Keyword(s):  
Structural Changes ◽  
Striated Muscle ◽  
Structural Basis ◽  
Ordered Structure ◽  
Compact Structure ◽  
Crude Homogenate ◽  
Myosin Filaments ◽  
The Cross ◽  
Cross Bridges ◽  
Muscle Myosin

We have used electron microscopy and proteolytic susceptibility to study the structural basis of myosin-linked regulation in synthetic filaments of scallop striated muscle myosin. Using papain as a probe of the structure of the head-rod junction, we find that this region of myosin is approximately five times more susceptible to proteolytic attack under activating (ATP/high Ca2+) or rigor (no ATP) conditions than under relaxing conditions (ATP/low Ca2+). A similar result was obtained with native myosin filaments in a crude homogenate of scallop muscle. Proteolytic susceptibility under conditions in which ADP or adenosine 5'-(beta, gamma-imidotriphosphate) (AMPPNP) replaced ATP was similar to that in the absence of nucleotide. Synthetic myosin filaments negatively stained under relaxing conditions showed a compact structure, in which the myosin cross-bridges were close to the filament backbone and well ordered, with a clear 14.5-nm axial repeat. Under activating or rigor conditions, the cross-bridges became clumped and disordered and frequently projected further from the filament backbone, as has been found with native filaments; when ADP or AMPPNP replaced ATP, the cross-bridges were also disordered. We conclude (a) that Ca2+ and ATP affect the affinity of the myosin cross-bridges for the filament backbone or for each other; (b) that the changes observed in the myosin filaments reflect a property of the myosin molecules alone, and are unlikely to be an artifact of negative staining; and (c) that the ordered structure occurs only in the relaxed state, requiring both the presence of hydrolyzed ATP on the myosin heads and the absence of Ca2+.

scholarly journals Structural changes accompanying phosphorylation of tarantula muscle myosin filaments.

1987 ◽  
Vol 105 (3) ◽  
pp. 1319-1327 ◽  
Author(s):  
R Craig ◽  
R Padrón ◽  
J Kendrick-Jones
Keyword(s):  
Gel Electrophoresis ◽  
Myosin Light Chain Kinase ◽  
Myosin Light Chain ◽  
Structural Changes ◽  
Striated Muscle ◽  
Light Chains ◽  
Regulatory Light Chains ◽  
Myosin Filaments ◽  
Muscle Myosin ◽  
Light Chains Of Myosin

Electron microscopy has been used to study the structural changes that occur in the myosin filaments of tarantula striated muscle when they are phosphorylated. Myosin filaments in muscle homogenates maintained in relaxing conditions (ATP, EGTA) are found to have nonphosphorylated regulatory light chains as shown by urea/glycerol gel electrophoresis and [32P]phosphate autoradiography. Negative staining reveals an ordered, helical arrangement of crossbridges in these filaments, in which the heads from axially neighboring myosin molecules appear to interact with each other. When the free Ca2+ concentration in a homogenate is raised to 10(-4) M, or when a Ca2+-insensitive myosin light chain kinase is added at low Ca2+ (10(-8) M), the regulatory light chains of myosin become rapidly phosphorylated. Phosphorylation is accompanied by potentiation of the actin activation of the myosin Mg-ATPase activity and by loss of order of the helical crossbridge arrangement characteristic of the relaxed filament. We suggest that in the relaxed state, when the regulatory light chains are not phosphorylated, the myosin heads are held down on the filament backbone by head-head interactions or by interactions of the heads with the filament backbone. Phosphorylation of the light chains may alter these interactions so that the crossbridges become more loosely associated with the filament backbone giving rise to the observed changes and facilitating crossbridge interaction with actin.

scholarly journals Recent X-ray Diffraction and Electron Microscope Studies of Striated Muscle

1967 ◽  
Vol 50 (6) ◽  
pp. 71-83 ◽  
Author(s):  
H. E. Huxley
Keyword(s):  
Active Sites ◽  
Striated Muscle ◽  
Structural Features ◽  
Actin Binding ◽  
X Ray Diffraction ◽  
X Ray ◽  
Thick Filaments ◽  
Myosin Filaments ◽  
The Cross ◽  
Cross Bridges

The sliding filament model for muscular contraction supposes that an appropriately directed force is developed between the actin and myosin filaments by some process in which the cross-bridges are involved. The cross-bridges between the filaments are believed to represent the parts of the myosin molecules which possess the active sites for ATPase activity and actin-binding ability, and project out sidewise from the backbone of the thick filaments. The arrangement of the cross-bridges is now being studied by improved low-angle X-ray diffraction techniques, which show that in a resting muscle, they are arranged approximately but not exactly in a helical pattern, and that there are other structural features of the thick filaments which give rise to additional long periodicities shown up by the X-ray diagram. The actin filaments also contain helically arranged subunits, and both the subunit repeat and the helical repeat are different from those in the myosin filaments. Diffraction diagrams can be obtained from muscles in rigor (when permanent attachment of the cross-bridges to the actin subunits takes place) and now, taking advantage of the great increase in the speed of recording, from actively contracting muscles. These show that changes in the arrangement of the cross-bridges are produced under both these conditions and are no doubt associated in contraction with the development of force. Thus configurational changes of the myosin component in muscle have been demonstrated: these take place without any significant over-all change in the length of the filaments.

scholarly journals Substructure and accessory proteins in scallop myosin filaments.

1989 ◽  
Vol 109 (2) ◽  
pp. 539-547 ◽  
Author(s):  
P Vibert ◽  
L Castellani
Keyword(s):  
Ionic Strength ◽  
Striated Muscle ◽  
Rotational Symmetry ◽  
Thick Filament ◽  
Accessory Proteins ◽  
Myosin Filaments ◽  
Cross Bridges ◽  
Low Ionic Strength ◽  
Muscle Myosin

Native myosin filaments from scallop striated muscle fray into subfilaments of approximately 100-A diameter when exposed to solutions of low ionic strength. The number of subfilaments appears to be five to seven (close to the sevenfold rotational symmetry of the native filament), and the subfilaments probably coil around one another. Synthetic filaments assembled from purified scallop myosin at roughly physiological ionic strength have diameters similar to those of native filaments, but are much longer. They too can be frayed into subfilaments at low ionic strength. Synthetic filaments share what may be an important regulatory property with native filaments: an order-disorder transition in the helical arrangement of myosin cross-bridges that is induced on activation by calcium, removal of nucleotide, or modification of a myosin head sulfhydryl. Some native filaments from scallop striated muscle carry short "end filaments" protruding from their tips, comparable to the structures associated with vertebrate striated muscle myosin filaments. Gell electrophoresis of scallop muscle homogenates reveals the presence of high molecular weight proteins that may include the invertebrate counterpart of titin, a component of the vertebrate end filament. Although the myosin molecule itself may contain much of the information required to direct its assembly, other factors acting in vivo, including interactions with accessory proteins, probably contribute to the assembly of a precisely defined thick filament during myofibrillogenesis.

scholarly journals LOCALIZATION OF MYOSIN FILAMENTS IN SMOOTH MUSCLE

1968 ◽  
Vol 37 (1) ◽  
pp. 105-116 ◽  
Author(s):  
Robert E. Kelly ◽  
Robert V. Rice
Keyword(s):  
Smooth Muscle ◽  
Cross Section ◽  
Striated Muscle ◽  
Thick Filament ◽  
Longitudinal Axis ◽  
Thin Filaments ◽  
Chicken Gizzard ◽  
Thick Filaments ◽  
Myosin Filaments ◽  
Muscle Myosin

Thick myosin filaments, in addition to actin filaments, were found in sections of glycerinated chicken gizzard smooth muscle when fixed at a pH below 6.6. The thick filaments were often grouped into bundles and run in the longitudinal axis of the smooth muscle cell. Each thick filament was surrounded by a number of thin filaments, giving the filament arrangement a rosette appearance in cross-section. The exact ratio of thick filaments to thin filaments could not be determined since most arrays were not so regular as those commonly found in striated muscle. Some rosettes had seven or eight thin filaments surrounding a single thick filament. Homogenates of smooth muscle of chicken gizzard also showed both thick and thin filaments when the isolation was carried out at a pH below 6.6, but only thin filaments were found at pH 7.4. No Z or M lines were observed in chicken gizzard muscle containing both thick and thin filaments. The lack of these organizing structures may allow smooth muscle myosin to disaggregate readily at pH 7.4.

scholarly journals Assembly of smooth muscle myosin into side-polar filaments.

1977 ◽  
Vol 75 (3) ◽  
pp. 990-996 ◽  
Author(s):  
R Craig ◽  
J Megerman
Keyword(s):  
Smooth Muscle ◽  
Opposite Polarity ◽  
Skeletal Muscle Myosin ◽  
Myosin Filaments ◽  
Skeletal Myosin ◽  
Cross Bridges ◽  
Bipolar Filament ◽  
Muscle Myosin

The in vitro assembly of myosin purified from calf aorta muscle has been studied by electron microscopy. Two types of filament are formed: short bipolar filament similar to those formed from skeletal muscle myosin, and longer "side-polar" filaments having cross bridges with a single polarity along the entire length of one side and the opposite polarity along the other side. Unlike the case with skeletal myosin filaments, antiparallel interactions between myosin molecules occur along the whole length of side-polar filaments. The side-polar structure may be related to the in vivo form of myosin in vertebrate smooth muscle.

scholarly journals Velocities of unloaded muscle filaments are not limited by drag forces imposed by myosin cross-bridges

2015 ◽  
Vol 112 (36) ◽  
pp. 11235-11240 ◽  
Author(s):  
Richard K. Brizendine ◽  
Diego B. Alcala ◽  
Michael S. Carter ◽  
Brian D. Haldeman ◽  
Kevin C. Facemyer ◽  
...  
Keyword(s):  
Duty Ratio ◽  
Step Size ◽  
Drag Forces ◽  
Contraction Speed ◽  
Myosin Filaments ◽  
Adp Release ◽  
Muscle Types ◽  
Cross Bridges ◽  
Muscle Myosin

It is not known which kinetic step in the acto-myosin ATPase cycle limits contraction speed in unloaded muscles (V0). Huxley’s 1957 model [Huxley AF (1957) Prog Biophys Biophys Chem 7:255–318] predicts that V0 is limited by the rate that myosin detaches from actin. However, this does not explain why, as observed by Bárány [Bárány M (1967) J Gen Physiol 50(6, Suppl):197–218], V0 is linearly correlated with the maximal actin-activated ATPase rate (vmax), which is limited by the rate that myosin attaches strongly to actin. We have observed smooth muscle myosin filaments of different length and head number (N) moving over surface-attached F-actin in vitro. Fitting filament velocities (V) vs. N to a detachment-limited model using the myosin step size d = 8 nm gave an ADP release rate 8.5-fold faster and ton (myosin’s attached time) and r (duty ratio) ∼10-fold lower than previously reported. In contrast, these data were accurately fit to an attachment-limited model, V = N·v·d, over the range of N found in all muscle types. At nonphysiologically high N, V = L/ton rather than d/ton, where L is related to the length of myosin’s subfragment 2. The attachment-limited model also fit well to the [ATP] dependence of V for myosin-rod cofilaments at three fixed N. Previously published V0 vs. vmax values for 24 different muscles were accurately fit to the attachment-limited model using widely accepted values for r and N, giving d = 11.1 nm. Therefore, in contrast with Huxley’s model, we conclude that V0 is limited by the actin–myosin attachment rate.

scholarly journals THE DOUBLE ARRAY OF FILAMENTS IN CROSS-STRIATED MUSCLE

1957 ◽  
Vol 3 (5) ◽  
pp. 631-648 ◽  
Author(s):  
H. E. Huxley
Keyword(s):  
Striated Muscle ◽  
The Other ◽  
Thin Sections ◽  
Alternative Models ◽  
Double Array ◽  
The Cross ◽  
Cross Bridges

The conditions under which one might expect to see the secondary filaments (if they exist) in longitudinal sections of striated muscle, are discussed. It is shown that these conditions were not satisfied in previously published works for the sections were too thick. When suitably thin sections are examined, the secondary filaments can be seen perfectly easily. It is also possible to see clearly other details of the structure, notably the cross-bridges between primary and secondary filaments, and the tapering of the primary filaments at their ends. The arrangement of the filaments and the changes associated with contraction and with stretch are identical to those already deduced from previous observations and described in terms of the interdigitating filament model in previous papers. There are therefore excellent grounds for believing that this model is correct. The alternative models which have been proposed appear to be incompatible both with the present observations and with much of the other available evidence.

scholarly journals Evidence from Insect Fibrillar Muscle about the Elementary Contractile Process

1967 ◽  
Vol 50 (6) ◽  
pp. 139-156 ◽  
Author(s):  
J. W. S. Pringle
Keyword(s):  
Ionic Strength ◽  
Atpase Activity ◽  
Striated Muscle ◽  
Contractile Activity ◽  
High Elasticity ◽  
Flight Muscles ◽  
The Cross ◽  
Cross Bridges ◽  
Low Ionic Strength ◽  
Water Bugs

Bundles of myofibrils prepared from the dorsal longitudinal flight muscles of giant water bugs show oscillatory contractile activity in solutions of low ionic strength containing ATP and 10-8-10-7 M Ca2+. This is due to delay between changes of length and changes of tension under activating conditions. The peculiarities of insect fibrillar muscle which give rise to this behavior are (1) the high elasticity of relaxed myofibrils, (2) a smaller degree of Ca2+ activation of ATPase activity in unstretched myofibrils and extracted actomyosin, and (3) a direct effect of stretch on ATPase activity. It is shown that the cross-bridges of striated muscle are probably formed from the heads of three myosin molecules and that in insect fibrillar muscle the cycles of mechanochemical energy conversion in the cross-bridges can be synchronized by imposed changes of length. This material is more suitable than vertebrate striated muscle for a study of the nature of the elementary contractile process.

scholarly journals Order-disorder structural transitions in synthetic filaments of fast and slow skeletal muscle myosins under relaxing and activating conditions.

2000 ◽  
Vol 47 (4) ◽  
pp. 1007-1017 ◽  
Author(s):  
Z A Podlubnaya ◽  
S L Malyshev ◽  
K Nieznański ◽  
D Stepkowski
Keyword(s):  
Skeletal Muscle ◽  
Ordered Structure ◽  
Structural Transitions ◽  
Fast Skeletal Muscle ◽  
Skeletal Muscle Myosin ◽  
Slow Skeletal Muscle ◽  
Random Arrangement ◽  
Vertebrate Muscle ◽  
Myosin Filaments ◽  
Muscle Myosin

In the previous study (Podlubnaya et al., 1999, J. Struc. Biol. 127, 1-15) Ca2+-induced reversible structural transitions in synthetic filaments of pure fast skeletal and cardiac muscle myosins were observed under rigor conditions (-Ca2+/+Ca2+). In the present work these studies have been extended to new more order-producing conditions (presence of ATP in the absence of Ca2+) aimed at arresting the relaxed structure in synthetic filaments of both fast and slow skeletal muscle myosin. Filaments were formed from column-purified myosins (rabbit fast skeletal muscle and rabbit slow skeletal semimebranosusproprius muscle). In the presence of 0.1 mM free Ca2+, 3 mM Mg2+ and 2 mM ATP (activating conditions) these filaments had a spread structure with a random arrangement of myosin heads and subfragments 2 protruding from the filament backbone. Such a structure is indistinguishable from the filament structures observed previously for fast skeletal, cardiac (see reference cited above) and smooth (Podlubnaya et al., 1999, J. Muscle Res. Cell Motil. 20, 547-554) muscle myosins in the presence of 0.1 mM free Ca2+. In the absence of Ca2+ and in the presence of ATP (relaxing conditions) the filaments of both studied myosins revealed a compact ordered structure. The fast skeletal muscle myosin filaments exhibited an axial periodicity of about 14.5 nm and which was much more pronounced than under rigor conditions in the absence of Ca2+ (see the first reference cited). The slow skeletal muscle myosin filaments differ slightly in their appearance from those of fast muscle as they exhibit mainly an axial repeat of about 43 nm while the 14.5 nm repeat is visible only in some regions. This may be a result of a slightly different structural properties of slow skeletal muscle myosin. We conclude that, like other filaments of vertebrate myosins, slow skeletal muscle myosin filaments also undergo the Ca2+-induced structural order-disorder transitions. It is very likely that all vertebrate muscle myosins possess such a property.

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