232 Ann. N.Y. Acad. Sci. 1047: 232–247 (2005). © 2005 New York Academy of Sciences. doi: 10.1196/annals.1341.021 Structure-Function Relation of the Myosin Motor in Striated Muscle MASSIMO RECONDITI, a,b MARCO LINARI, a,b LEONARDO LUCII, a,b ALEX STEWART, c YIN-BIAO SUN, d THEYENCHERI NARAYANAN, e TOM IRVING, f GABRIELLA PIAZZESI, a,b MALCOLM IRVING, d AND VINCENZO LOMBARDI a,b a Laboratorio di Fisiologia, DBAG, Università di Firenze, 50019 Sesto Fiorentino, Italy b Istituto Nazionale di Fisica della Materia, Operative Group in Grenoble, F-38043 Grenoble Cedex, France c Rosenstiel Center, Brandeis University, Waltham, Massachusetts 02545, USA d Randall Division of Cell and Molecular Biophysics, King’s College London, London SE1 1UL, UK e European Synchrotron Radiation Facility, F-38043 Grenoble Cedex, France f BioCAT Advanced Photon Source, Argonne National Laboratory, Argonne, Illinois 60439, USA ABSTRACT: Force and shortening in striated muscle are driven by a structural working stroke in the globular portion of the myosin molecules—the myosin head—that cross-links the myosin-containing filaments and the actin-containing filaments. We use time-resolved X-ray diffraction in single fibers from frog skeletal muscle to link the conformational changes in the myosin head deter- mined at atomic resolution in crystallographic studies with the kinetic and me- chanical features of the molecular motor in the preserved sarcomeric structure. Our approach exploits the improved brightness and collimation of the X-ray beams of the third generation synchrotrons by using X-ray interference between the two arrays of myosin heads in each bipolar myosin filament to measure with Å sensitivity the axial motions of myosin heads in situ during the synchronous execution of the working stroke elicited by rapid decreases in length or load imposed during an active isometric contraction. Changes in the intensity and interference-fine structure of the axial X-ray reflections following the mechanical perturbation allowed to establish the average conformation of the myosin heads during the active isometric contraction and the extent of tilt during the elastic response and during the subsequent working stroke. The myosin working stroke is 12 nm at low loads, which is consistent with crystal- lographic studies, while it is smaller and slower at higher loads. The load dependence of the size and speed of the myosin working stroke is the molecular determinant of the macroscopic performance and efficiency of muscle. KEYWORDS: myosin motor, striated muscle; working stroke; X-ray diffraction Address for correspondence: Prof. Vincenzo Lombardi, Laboratorio di Fisiologia, DBAG, Via G. Sansone 1, 50019 Sesto Fiorentino (FI), Italy. Voice: 39 055 457 2388; fax: 39 055 457 2387. vincenzo.lombardi@unifi.it