Conformations of Vertebrate Striated Muscle Myosin Monomers in Equilibrium with Filaments

Abstract

Porcine cardiac myosin monomers in equilibrium with filaments under physiological conditions were observed to have two conformations, extended and folded forms, upon electron microscopy and gel filtration HPLC. The conformational state was independent of ATP and the phosphorylation of regulatory light chain. The folded monomers of cardiac myosin were mainly in an open conformation with only one bend in the tail, and may not trap the hydrolysis products of ATP, as assessed by single turnover experiments. These properties are similar to those of the folded monomers of rabbit skeletal myosin [Katoh, T., Konishi, K., and Yazawa, M. (1998) J. Biol. Chem. 273, 11436-11439]. The conformational states of skeletal and cardiac myosin monomers were not affected by pH between 7.0 and 8.5. Although significant disassembly of filaments and thus an increase in the monomer concentration were observed with an increase in pH. The results indicate that the pH-dependent change in filament assembly is due to a shift of equilibrium between the filaments and extended monomers toward filament disassembly. The Mg2+-ATPase activity of these myosin monomers decreased with a decrease in the salt concentration below approximately 0.1 M, suggestive of the formation of a closed conformation similar to the conformation of 10S smooth myosin. The results suggest that the conformational change from the extended to the folded form is a common property of various myosin IIs.

The N-terminal Lobes of Both Regulatory Light Chains Interact with the Tail Domain in the 10 S-inhibited Conformation of Smooth Muscle Myosin

“…However, we can infer that regions c, d, e, and f (Fig. 5A) represent the major sites of tail-RLC interaction in 10 S, because they are consistent with images of single 10 S molecules (3,6,9,10,29). Region f overlaps with our previously identified BP-C108-RLC 10 S SMM cross-linked region (residues 1554 -1583, Fig.…”

“…Many isoforms of myosin II can adopt the 10 S conformation, including those from vertebrate smooth muscle (3,(5)(6)(7), molluscan striated (4,8), vertebrate striated (9), vertebrate cardiac (10), and vertebrate cytoplasmic (11)(12)(13)(14)(15)(16)(17)(18)(19) sources. Of these isoforms it appears that the regulated myosins (smooth, non-muscle, and molluscan) form the 10 S conformation to the greatest extent.…”

“…10 S has been detected in permeabilized embryonic, but not adult gizzard muscle (23), suggesting that the stability of filaments may be tissue-specific. Other motors such as myosin I (24), myosin V (10,25), and kinesin (26) can form a self-inhibited folded structures similar to 10 S myosin II. Therefore, the ability to adopt such folded structures may be an evolutionarily conserved property of motor proteins.…”

“…It is stabilized at physiological ionic strength in the presence of ATP. Electron micrographs of shadowed and negatively stained single molecules (3,5,9,10,29,30) show that the heads are bent down toward the tail and that the tail is bent at least twice into three regions of approximately equal length. The first bend is at the S2-LMM junction, and the second bending region appears to interact with the regulatory domains.…”

Three-dimensional structure of vertebrate cardiac muscle myosin filaments

“…The myosin heads appear to interact asymmetrically, with their motor domains pointing toward the filament bare zone. This conclusion is supported by the observation of similar asymmetric head-head interactions in isolated, negatively stained mouse cardiac myosin molecules under relaxing conditions (H. Jung and R.C., unpublished data), and by the observation of a folded conformation (suggestive of head-head interaction) in rotary shadowed cardiac myosin molecules (26). Attempts to dock the two-headed model (as well as individual ADP.Pi-like S1 molecules) in other orientations were unsuccessful.…”