RESUMEN
Myosins of fast and slow skeletal muscles differ by the isoform composition of the heavy and light chains. We compared functional characteristics of myosin from the fast (m. psoas) and slow (m. soleus) muscles of rabbits. The parameters of single actin-myosin interaction were measured in an optical trap, and the characteristics of the Ca2+ regulation of actin-myosin interaction were studied using an in vitro motility assay. The duration of interaction of myosin from the fast muscle with actin was shorter and the filament sliding velocity over this myosin was higher than the corresponding parameters for myosin from the slow muscle. The dependence pCa-velocity for myosin from the fast muscle was less sensitive to Ca2+ than that of slow muscle myosin. Thus, functional properties of myosin determine not only mechanical and kinetic characteristics of muscle contraction, but also the peculiarities of its Ca2+ regulation.
Asunto(s)
Fibras Musculares de Contracción Rápida/metabolismo , Fibras Musculares de Contracción Lenta/metabolismo , Miosinas/metabolismo , Actinas/metabolismo , Animales , Calcio/metabolismo , Contracción Muscular/fisiología , Pinzas Ópticas , ConejosRESUMEN
The results of the numerical simulation of the end-diastolic, end-systolic and stroke volumes of the left ventricle of the heart are presented. The simulation was based on a published simple kinetic model of cardiac muscle and approximation of the ventricle geometry with thick-wall cylinder where the fibre orientation varied linearly from sub-epicardium towards sub-endocardium. Blood flow was modelled with a liner compartment model. This simplified approach provides correct dependencies of the stroke volume on the pre- and afterload, namely end-diastolic pressure and peripheral resistance. The calculations show that the stroke volume is independent of arterial compliance and blood inertia.
Asunto(s)
Ventrículos Cardíacos/fisiopatología , Modelos Teóricos , Pericardio/fisiología , Presión Sanguínea/fisiología , Hemodinámica , Humanos , Contracción Miocárdica/fisiología , Volumen Sistólico/fisiologíaRESUMEN
The interaction of actin and myosin powers striated and smooth muscles and some other types of cell motility. Due to its highly ordered structure, skeletal muscle is a very convenient object for studying the general mechanism of the actin-myosin molecular motor. The history of investigation of the actin-myosin motor is briefly described. Modern concepts and data obtained with different techniques including protein crystallography, electron microscopy, biochemistry, and protein engineering are reviewed. Particular attention is given to X-ray diffraction studies of intact muscles and single muscle fibers with permeabilized membrane as they give insight into structural changes that underlie force generation and work production by the motor. Time-resolved low-angle X-ray diffraction on contracting muscle fibers using modern synchrotron radiation sources is used to follow movement of myosin heads with unique time and spatial resolution under near physiological conditions.
Asunto(s)
Actinas/fisiología , Músculo Esquelético/fisiología , Miosinas/fisiología , Citoesqueleto de Actina/química , Citoesqueleto de Actina/metabolismo , Citoesqueleto de Actina/fisiología , Actinas/química , Actinas/metabolismo , Animales , Humanos , Modelos Moleculares , Contracción Muscular , Músculo Esquelético/citología , Músculo Esquelético/metabolismo , Miofibrillas/química , Miofibrillas/metabolismo , Miofibrillas/fisiología , Miosinas/química , Miosinas/metabolismo , Unión Proteica , Conformación Proteica , Difracción de Rayos XRESUMEN
In the absence of adenosine triphosphate, the head domains of myosin cross-bridges in muscle bind to actin filaments in a rigor conformation that is expected to mimic that following the working stroke during active contraction. We used x-ray interference between the two head arrays in opposite halves of each myosin filament to determine the rigor head conformation in single fibers from frog skeletal muscle. During isometric contraction (force T(0)), the interference effect splits the M3 x-ray reflection from the axial repeat of the heads into two peaks with relative intensity (higher angle/lower angle peak) 0.76. In demembranated fibers in rigor at low force (<0.05 T(0)), the relative intensity was 4.0, showing that the center of mass of the heads had moved 4.5 nm closer to the midpoint of the myosin filament. When rigor fibers were stretched, increasing the force to 0.55 T(0), the heads' center of mass moved back by 1.1-1.6 nm. These motions can be explained by tilting of the light chain domain of the head so that the mean angle between the Cys(707)-Lys(843) vector and the filament axis increases by approximately 36 degrees between isometric contraction and low-force rigor, and decreases by 7-10 degrees when the rigor fiber is stretched to 0.55 T(0).
Asunto(s)
Biomimética/métodos , Cristalografía por Rayos X/métodos , Contracción Isométrica , Proteínas Motoras Moleculares/química , Movimiento , Músculo Esquelético/fisiopatología , Miosinas/química , Rigor Mortis/fisiopatología , Actinas/química , Actinas/ultraestructura , Animales , Elasticidad , Modelos Biológicos , Modelos Moleculares , Proteínas Motoras Moleculares/ultraestructura , Músculo Esquelético/ultraestructura , Miosinas/ultraestructura , Conformación Proteica , Rigor Mortis/patología , Estrés Mecánico , Relación Estructura-ActividadRESUMEN
Generation of force and shortening in striated muscle is due to the cyclic interactions of the globular portion (the head) of the myosin molecule, extending from the thick filament, with the actin filament. The work produced in each interaction is due to a conformational change (the working stroke) driven by the hydrolysis of ATP on the catalytic site of the myosin head. However, the precise mechanism and the size of the force and length step generated in one interaction are still under question. Here we reinvestigate the endothermic nature of the force-generating process by precisely determining, in tetanized intact frog muscle fibres under sarcomere length control, the effect of temperature on both isometric force and force response to length changes. We show that raising the temperature: (1) increases the force and the strain of the myosin heads attached in the isometric contraction by the same amount (approximately 70 %, from 2 to 17 degrees C); (2) increases the rate of quick force recovery following small length steps (range between -3 and 2 nm (half-sarcomere)-1) with a Q10 (between 2 and 12 degrees C) of 1.9 (releases) and 2.3 (stretches); (3) does not affect the maximum extent of filament sliding accounted for by the working stroke in the attached heads (10 nm (half-sarcomere)-1). These results indicate that in isometric conditions the structural change leading to force generation in the attached myosin heads can be modulated by temperature at the expense of the structural change responsible for the working stroke that drives filament sliding. The energy stored in the elasticity of the attached myosin heads at the plateau of the isometric tetanus increases with temperature, but even at high temperature this energy is only a fraction of the mechanical energy released by attached heads during filament sliding.
Asunto(s)
Contracción Isométrica/fisiología , Modelos Biológicos , Fibras Musculares Esqueléticas/fisiología , Músculo Esquelético/fisiología , Temperatura , Animales , Elasticidad , Músculo Esquelético/citología , Miosinas/fisiología , Rana esculenta , TermodinámicaRESUMEN
Axial x-ray diffraction patterns from single intact fibers of frog skeletal muscle were recorded by using a highly collimated x-ray beam at the European Synchrotron Radiation Facility. During isometric contraction at sarcomere lengths 2.2-3.2 microm, the M3 x-ray reflection, associated with the repeat of myosin heads along the filaments, was resolved into two peaks. The total M3 intensity decreased linearly with increasing sarcomere length and was directly proportional to the degree of overlap between myosin and actin filaments, showing that it comes from myosin heads in the overlap region. The separation between the M3 peaks was smaller at longer sarcomere length and was quantitatively explained by x-ray interference between myosin heads in the two overlap regions of each sarcomere. The relative intensity of the M3 peaks was independent of sarcomere length, showing that the axial periodicities of the nonoverlap and overlap regions of the myosin filament have the same value, 14.57 nm, during active contraction. In resting fibers the periodicity is 14.34 nm, so muscle activation produces a change in myosin filament structure in the nonoverlap as well as the overlap part of the filament. The results establish x-ray interferometry as a new tool for studying the motions of myosin heads during muscle contraction with unprecedented spatial resolution.
Asunto(s)
Músculo Esquelético/química , Músculo Esquelético/ultraestructura , Sarcómeros/química , Sarcómeros/ultraestructura , Animales , Contracción Muscular , Rana temporaria , Difracción de Rayos XRESUMEN
Muscle contraction is driven by a change in shape of the myosin head region that links the actin and myosin filaments. Tilting of the light-chain domain of the head with respect to its actin-bound catalytic domain is thought to be coupled to the ATPase cycle. Here, using X-ray diffraction and mechanical data from isolated muscle fibres, we characterize an elastic bending of the heads that is independent of the presence of ATP. Together, the tilting and bending motions can explain force generation in isometric muscle, when filament sliding is prevented. The elastic strain in the head is 2.0-2.7 nm under these conditions, contributing 40-50% of the compliance of the muscle sarcomere. We present an atomic model for changes in head conformation that accurately reproduces the changes in the X-ray diffraction pattern seen when rapid length changes are applied to muscle fibres both in active contraction and in the absence of ATP. The model predictions are relatively independent of which parts of the head are assumed to bend or tilt, but depend critically on the measured values of filament sliding and elastic strain.
Asunto(s)
Contracción Muscular/fisiología , Miosinas/fisiología , Actinas/química , Actinas/fisiología , Adenosina Trifosfato/fisiología , Animales , Elasticidad , Proteínas Motoras Moleculares , Fibras Musculares Esqueléticas/fisiología , Miosinas/química , Conformación Proteica , Rana temporaria , Difracción de Rayos XRESUMEN
Step changes in length (between -3 and +5 nm per half-sarcomere) were imposed on isolated muscle fibers at the plateau of an isometric tetanus (tension T0) and on the same fibers in rigor after permeabilization of the sarcolemma, to determine stiffness of the half-sarcomere in the two conditions. To identify the contribution of actin filaments to the total half-sarcomere compliance (C), measurements were made at sarcomere lengths between 2.00 and 2.15 microm, where the number of myosin cross-bridges in the region of overlap between the myosin filament and the actin filament remains constant, and only the length of the nonoverlapped region of the actin filament changes with sarcomere length. At 2.1 microm sarcomere length, C was 3.9 nm T0(-1) in active isometric contraction and 2.6 nm T0(-1) in rigor. The actin filament compliance, estimated from the slope of the relation between C and sarcomere length, was 2.3 nm microm(-1) T0(-1). Recent x-ray diffraction experiments suggest that the myosin filament compliance is 1.3 nm microm(-1) T0(-1). With these values for filament compliance, the difference in half-sarcomere compliance between isometric contraction and rigor indicates that the fraction of myosin cross-bridges attached to actin in isometric contraction is not larger than 0.43, assuming that cross-bridge elasticity is the same in isometric contraction and rigor.
Asunto(s)
Actinas/fisiología , Contracción Isométrica/fisiología , Fibras Musculares Esqueléticas/fisiología , Músculo Esquelético/fisiología , Miosinas/fisiología , Sarcómeros/fisiología , Actinas/química , Animales , Elasticidad , Técnicas In Vitro , Cinética , Modelos Biológicos , Relajación Muscular , Miosinas/química , Unión Proteica , Rana esculenta , Factores de TiempoRESUMEN
Muscle contraction is generally thought to involve tilting of the light chain region of the myosin head. This could account for 5-10 nm of axial displacement as it moves from nearly perpendicular to the filament axis (the state at the beginning of the working stroke) to the rigor conformation (at the end of the working stroke). According to the kinetic model of Huxley and Simmons, the extent that a cross-bridge progresses through the working stroke depends on the mechanical conditions. A large tilting occurs only when the fibre is allowed to shorten. Evidence for such tilting was provided by the changes in intensity of the third myosin meridional reflection (M3) following a step release. However, there is little change in the M3 intensity when a force increase is elicited by a 10 degrees C temperature jump, and these results were interpreted to indicate that tilting is not the structural transition responsible for force generation. Here we present a simulation of the changes in the intensity of the M3 reflection elicited by step changes in either length or temperature, based on the atomic model of the actin-myosin head complex. The results show that the same set of assumptions for the motions associated with the working stroke can predict the response to both kinds of perturbation. The main difference is due to the larger extent of the working stroke elicited by the length step.