Atomic model of the human cardiac muscle myosin filament

Abstract

Of all the myosin filaments in muscle, the most important in terms of human health, and so far the least studied, are those in the human heart. Here we report a 3D single-particle analysis of electron micrograph images of negatively stained myosin filaments isolated from human cardiac muscle in the normal (undiseased) relaxed state. The resulting 28-Å resolution 3D reconstruction shows axial and azimuthal (no radial) myosin head perturbations within the 429-Å axial repeat, with rotations between successive 132 Å-, 148 Å-, and 149 Å-spaced crowns of heads close to 60°, 35°, and 25°(all would be 40°in an unperturbed three-stranded helix). We have defined the myosin head atomic arrangements within the three crown levels and have modeled the organization of myosin subfragment 2 and the possible locations of the 39 Å-spaced domains of titin and the cardiac isoform of myosin-binding protein-C on the surface of the myosin filament backbone. Best fits were obtained with head conformations on all crowns close to the structure of the two-headed myosin molecule of vertebrate chicken smooth muscle in the dephosphorylated relaxed state. Individual crowns show differences in head-pair tilts and subfragment 2 orientations, which, together with the observed perturbations, result in different intercrown head interactions, including one not reported before. Analysis of the interactions between the myosin heads, the cardiac isoform of myosin-binding protein-C, and titin will aid in understanding of the structural effects of mutations in these proteins known to be associated with human cardiomyopathies.thick filaments 3D structure | human heart muscle | electron microscopy | image processing M uscle contraction involves the interaction between actin and myosin filaments that leads to force production mediated by the hydrolysis of ATP. Myosin filaments are assemblies of myosin molecules and accessory proteins. Myosin molecules consist of two heavy chains arranged to form two globular domains [two subfragment-1s (S1s)] at the N-terminal end and a long rod-like α-helical coiled-coil domain (the myosin tail). Each of the globular domains with its associated essential and regulatory light chains forms a myosin head containing the ATPase activity necessary for contraction (1). This is linked to the N-terminal part of the tail known as the subfragment-2 (S2) region. Myosin filaments are generally bipolar, with the myosin tails packing together in an almost cylindrical backbone and the heads projecting out laterally in a helical or quasihelical fashion at intervals of ∼143 Å. In the myosin filaments of vertebrate striated muscles, the myosin heads are arranged in a threestranded quasihelical array on the filament surface (2) with the accessory proteins titin (3, 4), myosin-binding protein-C (MyBP-C) (5), and possibly the MyBP-C analog, X protein (6), located on the surface of the backbone. MyBP-C is located within the C zones of the myosin filament, which are regions ∼3,500 Å long centrally located in the two halves of the bipolar filament...

The myosin mesa and the basis of hypercontractility caused by hypertrophic cardiomyopathy mutations

“…Structural evidence for interaction of S1 with the proximal S2 tail from the same molecule has been observed for isolated heavy meromyosin (HMM, a soluble fragment of myosin consisting of two S1 heads attached to S2) 39–42 , myosin molecules 40–43 , and intact thick filaments 1–3,7,8 . The first 3D structure of a folded state of myosin, named the interacting heads motif (IHM), came from a cryo-EM study of unphosphorylated smooth muscle myosin 39 and describes a 2.0-nm-resolution map of an asymmetric interaction between the S1 domains.…”

“…The IHM has also been observed in other classes of muscle myosin, including striated muscle myosins 41 , and in images of intact thick filaments isolated from a variety of muscle types 1–6 . Thick-filament structures of 2.0- to 4.0-nm resolution have been obtained with cryo-EM for tarantula skeletal muscle 1,2 , Limulus telson muscle 3 , and vertebrate cardiac muscle 7,8 . The solved crystal structure of human β-cardiac proximal S2 reveals possible charge interactions between the S2 and the S1 (ref.…”

Deletion of the titin N2B region accelerates myofibrillar force development but does not alter relaxation kinetics

“…The net effect of how coexisting wild-type and N2B-deficient titin isoforms will affect passive stiffness is difficult to predict, especially as data on the spatial organization of titin within the intact sarcomere structure is scarce. Based on three-dimensional single-particle analysis of electron-microscopy micrographs, Aband titin might exist in pairs (Al Khayat et al, 2013;Zoghbi et al, 2008). The distal tandem Ig segment has been suggested to form a higher-order structure of a side-to-side hexamer (Houmeida, et al, 2008).…”

“…Furthermore, a recent X-ray diffraction study performed by Farman and co-workers showed that LDA might be based on the ordering of weakly bound cross-bridge orientation prior to activation (Farman et al, 2011). Titin winds along the myosin filament (Al Khayat et al, 2013), and it is tempting to speculate that increased passive tension affects myosin such that it enhances the probability of cross-bridge transitions to forcegenerating states, as reflected by the higher k ACT and k TR in myofibrils from knockout compared with wild-type mice in our study. Regarding the physiological situation when the sarcomeres are stretched during diastolic filling of the heart, the acceleration of cross-bridge cycling by increased titin-based passive tension might compensate for the potential slowing down of cross-bridge cycling resulting from the reduction of filament overlap.…”

Targeting the sarcomere to correct muscle function

“…The S2 and light meromyosin (LMM) subfragments comprise a long tail that dimerizes to form a coiled coil. Polymerization of LMM produces a rod-like thick filament, with soluble S2 pointing radially outwards 142,143 . The S1 subfragment comprises the globular multi-domain head, which binds actin and hydrolyses ATP, and the neck region, which is stabilized by the myosin essential light chain (ELC) and the regulatory light chain (RLC) 144 .…”