Soft Report - Dipartimento di Fisica - Sapienza

Scientific Report – Non Equilibrium Dynamics and ComplexityThe Molecular Mechanism of Muscle ContractionThe cells of the striated muscle, called fibres, areconstituted by ca 2 µm long elementary units, thesarcomeres, that repeat along the axis of the fibre(Fig. 1). In each half-sarcomere, thick (myosincontaining)filaments originating from the M line atthe centre of the sarcomere partially overlap withthin (actin-containing) filaments originating from theZ line bounding the sarcomere. During musclecontraction the generation of the force that pulls theactin filament towards the centre of the sarcomere isdue to a structural working stroke in the globularhead of the myosin cross-linking the myosin and theactin filaments. The work produced is accounted forby the hydrolysis of ATP on the catalytic site of themyosin head. Despite the mass of information frommechanical, biochemical and energetic studies thegap between cellular and molecular levels ofdescription of the myosin motor remains large.Protein crystallography has provided a model of themyosin working stroke with atomic resolution.However, the function of myosin in situ depends onthe interaction between conformational changes inthe motor protein and external force or motion, andthis cannot be reproduced in crystallographic studies.In isolated intact cells from frog muscle, myosinmotors can be synchronised by length or force stepscontrolled at half-sarcomere level and the relatedstructural changes can be recorded with timeresolvedsmall angle X-ray diffraction (SAXS) usingsynchrotron light. The brightest axial reflection of thediffraction pattern from single fibres, called M3,originates from the 14.5 nm axial repeat of themyosin motors along the filament axis (Fig. 1) and issensitive to axial movements of the myosin headsduring the working stroke [1].A breakthrough for SAXS technique has been thefinding that with the spatial resolution of 3rdgeneration synchrotrons (ESRF, Grenoble, France;APS, Argonne, IL, USA) it is possible to record thefringes generated in the M3 reflection by theinterference between the two arrays of myosinmotors in each sarcomere [2]. Due to the bipolararrangement of the myosin motors in the two halvesof the sarcomere, the interference effect provides Åscale direct measure of the axial movement of themotors [3]. The changes in interference fringes ofthe M3 reflection, following stepwise reduction of theZ-line14.5 nmMyosin filamentMyosinheadsM-lineActin filamentFig. 1: Structural model of muscle contraction atthe level of the sarcomere. Arrangement of the actinand myosin molecules in the muscle sarcomere.Grey, actin filament; blue, myosin filament; red;myosin heads. Sarcomere shortening (transitionfrom upper to lower panel) is associated with tiltingof myosin heads so that actin filaments are pulledtoward the M-line.1.00.50.00-5-10-15-20abForce ( T 0 units)L 0 1L 2s 2L 2e L 33Length change (nm hs -1 )0 5 10 15 20Time (ms)0.066 0.068 0.070 0.072Fig. 2: Mechanical and structural responses to aload step. (a) Load step normalised by theisometric force T 0. (b) Length change in nm perhalf-sarcomere; numbers next to the recordindicate the various phases of the shortening: theelastic change in strain (1); the early sliding due tothe synchronised working stroke in the myosinheads (2); the pause (3) and steady sliding (4) dueto detachment/attachment of myosin heads fartheralong the actin filament. (c) Axial intensitydistribution in the region of the M3 reflection at theperiods corresponding to the X-ray exposure timesshown in (b): brown L 0, isometric contraction;orange L 2s, start of phase 2; pink L 2e, end of phase2; blue L 3, end of phase 3. Myosin heads movetowards the centre of the sarcomere during phase 2and detach from actin during phase 3.force from the isometric value with a force feedbackcontrol, showed that the myosin working stroke is 11nm and takes