Myosin SI fragment

Figure 14.15 Stmcture of the SI fragment of chicken myosin as a Richardson diagram (a) and a space-filling model (b). The two light chains are shown in magenta and yellow. The heavy chain is colored according to three proteolytic fragments produced by trypsin a 25-kDa N-terminal domain (green) a central 50-kDa fragment (red) divided by a cleft into a 50K upper and a 50K lower domain and a 20-kDa C-terminal domain (blue) that links the myosin head to the coiled-coil tail. The 50-kDa and 20-kDa domains both bind actin, while the 25-kDa domain binds ATP. [(b) Courtesy of 1. Rayment.]...

Figure 4.5. Structure of myosin. Myosin comprises both light and heavy chains. The heavy chains may be cleaved by trypsin to generate light meromyosin (LMM) and heavy mero-myosin (HMM). Papain digestion of HMM yields subfragments SI and S2 each SI fragment contains an ATPase site and an actin-binding site. The light chains modify the activity of the ATPase.

Using film an,d delay-line detectors on the DESY and XI1 benches, a study of the binding of exogeneous myosin heads (SI fragments) to insect fli t muscle in the rigor... 

Figure 34.3. Myosin Dissection. Treatment of muscle myosin with proteases forms stable fragments, including subfragments SI and S2 and light meromyosin. Each SI fragment includes the head (shown in yellow and pink) from the heavy chain and one copy of each light chain (shown in blue and orange).

Figure 34.4. Myosin Structure at High Resolution. The structure of the SI fragment from muscle myosin reveals the presence of a P-loop NTPase domain (shaded in purple). An a helix that extends from this domain is the binding site for the two light chains.

Figure 34.10. Lever-Arm Motion. Two forms of the SI fragment of scallop muscle myosin. Dramatic conformational changes are observed when the identity of the bound nucleotide changes from ADP-V043- to ADP or vice versa, including a nearly 90-degree reorientation of the lever arm.

Let us first consider the structure of myosin. The results of electron microscopic studies of skeletal-muscle myosin show it to be a two-headed structure linked to a long stalk (Figure 34.2). As we saw in Chapter 33, limited proteolysis can be a powerful tool in probing the activity of large proteins. The treatment of myosin with trypsin and papain results in the formation of four fragments two SI fragments an S2 fragment, also called heavy... 

Figure 12.9 Single moLecuLe studies can reveal enzyme kinetic information. Left the acto-myosin ATPase cycle lined up with a typical binding event the event begins when myosin binds to actin and ends when an ATP molecule diffuses into the binding cleft, causing it to dissociate from the actin filament. Right the event lifetimes of rabbit skeletal myosin-II SI fragment are stochastic and show an exponential distribution because each event is terminated by a single Poisson process (ATP binding). Note that the inset has two points removed, as zeros cannot be plotted on a log scale...

Cleavage of the -850-residue SI heads with trypsin yields mainly three large fragments that correspond to structural domains of the intact protein as shown in Fig. 19-15. They are known as the 25-kDa (N-terminal), 50-kDa, and 20-kDa fragments, and for myosin from D. discoideum correspond to residues 1 to 204,216 to 626, and 647 to 843, respectively. The ATP-binding site is in a deep cleft between the 20-kDa and 50-kDa... 

Structure of the myosin heads. Myosin and myosin fragments can be isolated in large quantities, but they have been difficult to crystallize. However, Payment and coworkers purified SI heads cleaved from chicken myosin by papain and subjected them to reductive methylation (using a dimethylamine-borane complex see also Eq. 3-34). With most of the lysine side chain amino groups converted to dimethylamine groups, high-quality crystals were obtained, and a structure was determined by X-ray diffraction. ... 

Other single molecule techniques involve direct observation of motor molecules or of SI myosin fragments tagged with highly fluorescent labels. ... 

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