Structural Characterization of the Binding of Myosinmiddle dotADPmiddle dotPi to Actin in Permeabilized Rabbit Psoas Muscle

@article{citeulike:1365812, abstract = {When myosin is attached to actin in a muscle cell, various structures in the filaments are formed. The two strongly bound states (A{middle dot}M{middle dot}ADP and A{middle dot}M) and the weakly bound A{middle dot}M{middle dot}ATP states are reasonably well understood. The orientation of the strongly bound myosin heads is uniform ("stereospecific" attachment), and the attached heads exhibit little spatial fluctuation. In the prehydrolysis weakly bound A{middle dot}M{middle dot}ATP state, the orientations of the attached myosin heads assume a wide range of azimuthal and axial angles, indicating considerable flexibility in the myosin head. The structure of the other weakly bound state, A{middle dot}M{middle dot}ADP{middle dot}Pi, however, is poorly understood. This state is thought to be the critical pre-power-stroke state, poised to make the transition to the strongly binding, force-generating states, and hence it is of particular interest for understanding the mechanism of contraction. However, because of the low affinity between myosin and actin in the A{middle dot}M{middle dot}ADP{middle dot}Pi state, the structure of this state has eluded determination both in isolated form and in muscle cells. With the knowledge recently gained in the structures of the weakly binding M{middle dot}ATP, M{middle dot}ADP{middle dot}Pi states and the weakly attached A{middle dot}M{middle dot}ATP state in muscle fibers, it is now feasible to delineate the in vivo structure of the attached state of A{middle dot}M{middle dot}ADP{middle dot}Pi. The series of experiments presented in this article were carried out under relaxing conditions at 25{degrees}C, where [~]95\% of the myosin heads in the skinned rabbit psoas muscle contain the hydrolysis products. The affinity for actin is enhanced by adding polyethylene glycol (PEG) or by lowering the ionic strength in the bathing solution. Solution kinetics and binding constants were determined in the presence and in the absence of PEG. When the binding between actin and myosin was increased, both the myosin layer lines and the actin layer lines increased in intensity, but the intensity profiles did not change. The configuration (mode) of attachment in the A{middle dot}M{middle dot}ADP{middle dot}Pi state is thus unique among the intermediate attached states of the cross-bridge ATP hydrolysis cycle. One of the simplest explanations is that both myosin filaments and actin filaments are stabilized (e.g., undergo reduced spatial fluctuations) by the attachment. The alignment of the myosin heads in the thick filaments and the alignment of the actin monomers in the thin filaments are improved as a result. The compact atomic structure of M{middle dot}ADP{middle dot}Pi with strongly coupled domains may contribute to the unique attachment configuration: the "primed" myosin heads may function as "transient struts" when attached to the thin filaments. 10.1529/biophysj.106.086918}, author = {Xu, Sengen and Gu, Jin and Belknap, Betty and White, Howard and Yu, Leepo C. }, citeulike-article-id = {1365812}, doi = {10.1529/biophysj.106.086918}, journal = {Biophys. J.}, keywords = {experiment, fiber, motor, muscle, myosin, xray-diffraction}, month = {November}, number = {9}, pages = {3370--3382}, posted-at = {2007-06-05 16:53:04}, priority = {2}, title = {Structural Characterization of the Binding of Myosin{middle dot}ADP{middle dot}Pi to Actin in Permeabilized Rabbit Psoas Muscle}, url = {http://dx.doi.org/10.1529/biophysj.106.086918}, volume = {91}, year = {2006} }

TY - JOUR ID - citeulike:1365812 L3 - citeulike-article-id:1365812 TI - Structural Characterization of the Binding of Myosin{middle dot}ADP{middle dot}Pi to Actin in Permeabilized Rabbit Psoas Muscle JF - Biophys. J. VL - 91 IS - 9 SP - 3370 EP - 3382 N2 - When myosin is attached to actin in a muscle cell, various structures in the filaments are formed. The two strongly bound states (A{middle dot}M{middle dot}ADP and A{middle dot}M) and the weakly bound A{middle dot}M{middle dot}ATP states are reasonably well understood. The orientation of the strongly bound myosin heads is uniform ("stereospecific" attachment), and the attached heads exhibit little spatial fluctuation. In the prehydrolysis weakly bound A{middle dot}M{middle dot}ATP state, the orientations of the attached myosin heads assume a wide range of azimuthal and axial angles, indicating considerable flexibility in the myosin head. The structure of the other weakly bound state, A{middle dot}M{middle dot}ADP{middle dot}Pi, however, is poorly understood. This state is thought to be the critical pre-power-stroke state, poised to make the transition to the strongly binding, force-generating states, and hence it is of particular interest for understanding the mechanism of contraction. However, because of the low affinity between myosin and actin in the A{middle dot}M{middle dot}ADP{middle dot}Pi state, the structure of this state has eluded determination both in isolated form and in muscle cells. With the knowledge recently gained in the structures of the weakly binding M{middle dot}ATP, M{middle dot}ADP{middle dot}Pi states and the weakly attached A{middle dot}M{middle dot}ATP state in muscle fibers, it is now feasible to delineate the in vivo structure of the attached state of A{middle dot}M{middle dot}ADP{middle dot}Pi. The series of experiments presented in this article were carried out under relaxing conditions at 25{degrees}C, where [~]95% of the myosin heads in the skinned rabbit psoas muscle contain the hydrolysis products. The affinity for actin is enhanced by adding polyethylene glycol (PEG) or by lowering the ionic strength in the bathing solution. Solution kinetics and binding constants were determined in the presence and in the absence of PEG. When the binding between actin and myosin was increased, both the myosin layer lines and the actin layer lines increased in intensity, but the intensity profiles did not change. The configuration (mode) of attachment in the A{middle dot}M{middle dot}ADP{middle dot}Pi state is thus unique among the intermediate attached states of the cross-bridge ATP hydrolysis cycle. One of the simplest explanations is that both myosin filaments and actin filaments are stabilized (e.g., undergo reduced spatial fluctuations) by the attachment. The alignment of the myosin heads in the thick filaments and the alignment of the actin monomers in the thin filaments are improved as a result. The compact atomic structure of M{middle dot}ADP{middle dot}Pi with strongly coupled domains may contribute to the unique attachment configuration: the "primed" myosin heads may function as "transient struts" when attached to the thin filaments. 10.1529/biophysj.106.086918 KW - experiment KW - fiber KW - motor KW - muscle KW - myosin KW - xray-diffraction AU - Xu, Sengen AU - Gu, Jin AU - Belknap, Betty AU - White, Howard AU - Yu, Leepo PY - 2006/11/01/ UR - http://dx.doi.org/10.1529/biophysj.106.086918 ER -