Helix relay

F. Twisting the /3-Sheet Takes the Kink Out of the Relay Helix. 173... 

State SW1 Outer Cleft Inner Cleft SW2 / -sheet P-loop Relay Helix Converter- Lever... 

Fig. 3. Details of the seven-stranded /1-sheet and associated structures (A and B) in the post-rigor conformation and (C and D) in the pre-powerstroke conformation. The orientation of A and C is at right angles to that shown in Fig. 2. When attached to actin, it corresponds to that shown in Fig. 5B. The colors are as in Fig. 2. The views shown in B and D are at right angles to A and C looking out radially from the axis of the actin helix. Note the kink in the relay helix shown in C and D that leads to a 60° rotation of the converter domain. This in turn rotates the lever arm 60°. The P-loop (which constitutes the ATP-binding site) and the adjoining a-helix are shown in yellow. The flanking switch sequences (1 and 2) are also shown. The strands of the /1-sheet are numbered from the N-terminal (distal) end of the sheet. The lower part of strand 5 (light blue) constitutes switch 2. In the post-rigor state, switch 2 lies out of the plane of the /1-sheet (open) and in the pre-powerstroke state switch 2 is in the plane of the /1-sheet (closed).

It is important to note that the binding of ATP places the /P, into the pocket formed by SW1 and the P-loop, which promotes the moving in of SW2. The effects of this movement are twofold the relay helix is kinked and thereby re-primed, and the enzymatically active site is formed. In the pre-powerstroke state, the nucleotide is completely enclosed by the positioning of SW1 and SW2. This conformation appears to be the enzymatically active form of myosin. Moreover, this is the preferred stable structure of myosin in the presence of nucleotide. 

The position of the converter domain depends on whether the relay helix has a kink near its middle point. The kink in the relay helix occurs in the pre-powerstroke state. The kink leads to a rotation of the converter domain through 60°. Removing the kink causes the lever arm to rotate back by 60°, which is the elementary structural event in the powerstroke. Conversely, creating the kink is the priming action necessary to reach the start of the powerstroke. 

Myosin therefore appears to possess two mechanisms for relieving the kink in the relay helix either by moving out the lower 50K domain (SW2 closed to SW2 open) or by twisting the /1-sheet, which gives the distal end of the relay helix more space. The first mechanism (in the reverse order, SW2 open to SW2 closed) occurs in the absence of actin after ATP binds in the nucleotide pocket. This is the recovery stroke leading to the priming of the powerstroke. However, when the myosin is bound to actin, it appears that only the second mechanism is relevant (indeed, it would be rather... 

Fig. 6. The /Tsheet can twist. A comparison of the pre-powerstroke state (A) with the rigor-like state (B). For this comparison, strands 6 and 7 of the /i-sheet (shown edge on) have been aligned (this corresponds closely with an alignment of the lower 50K domains). The SW2 elements (light blue, below strand 5) in the two states are in very similar positions, as is the proximal part of the relay helix (light blue). In A, the outer part of the relay helix (dark blue) is kinked. In B, the relay helix is straight. A comparison of A and B shows that the distal end of the /l-sheet (strands 1-4) twists through 7-8°. Note that this leads to a displacement of the P-loop.

Figure 34.11. Relay Helix. A superposition of key elements in two forms of scallop myosin reveals the structural changes that are transmitted by the relay helix from the switch I and switch II loops to the base of the lever arm. The switch I and switch II loops interact with V043- in the position that would be occupied by the y-phosphate group of ATP. The structure of the ADP - V043- niyosin complex is shown in lighter colors.

This conformational change allows a long a helix (termed the relay helix) to adjust its position. The carboxyl-terminal end of the relay helix interacts with struc-tures at the base of the lever arm, and so a change in the position of the relay helix leads to a reorientation of the lever arm. 

Analogous conformational changes take place in ki-iiesin. The kinesins also have a relay helix that can adopt different configurations when kinesin binds different nucleotides. Kinesin lacks an a-helical lever arm, however. 

Describe the large conformational difference between myosin-ADP and myosin-ATP (same as myosin-ADP-vanadate). Locate switch 1, switch 11, and the relay helix in relation to the P-loop, and explain how they cause the protein to flex. 

Both proteins flex dramatically when ATP binds to the P-loop area. The change in myosin is amplified by nearby structures like the relay helix, and by the length of the lever arm, but the change is generally parallel. Adenylate kinase maintains an equilibrium between ATP and ADP and must be extremely ancient. Myosin is confined to eukaryotes, and hence should be somewhat younger. 

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