Shaking the myosin family tree: biochemical kinetics defines four types of myosin motor

Abstract

Although all myosin motors follow the same basic cross-bridge cycle, they display a large variety in the rates of transition between different states in the cycle, allowing each myosin to be finely tuned for a specific task. Traditionally, myosins have been classified by sequence analysis into a large number of sub-families (∼35). Here we use a different method to classify the myosin family members which is based on biochemical and mechanical properties. The key properties that define the type of mechanical activity of the motor are duty ratio (defined as the fraction of the time myosin remains attached to actin during each cycle), thermodynamic coupling of actin and nucleotide binding to myosin and the degree of strain-sensitivity of the ADP release step. Based on these properties we propose to classify myosins into four different groups: (I) fast movers, (II) slow/efficient force holders, (III) strain sensors and (IV) gates.

Fig 1. The ATP driven actin-myosin cross-bridge cycle

The myosin cross-bridge is shown as consisting of 3 major parts. 1. The central core of the myosin head is shown in red (filled circle) with the lower 50k domain projecting up to make contact with one actin monomer (grey circles). The lower 50k domain is one half of the major cleft that splits the actin binding site. The P-loop that binds to the γPi of ATP is part of the central core. This red segment is drawn as a fixed reference point during the cycle. 2. The upper 50 K domain with SW-1 (switch 1) is shown in yellow. This is drawn as two parts; the upper jaw of the major cleft that splits the actin binding site and a yellow ring with SW-1 projecting into the center. 3. A blue ring represents the relay loop with SW-2 projecting to the center while the converter domain and the lever arm (shown with 2 light chains attached) is projected out from the ring. The nucleotide pocket is shown as lying on top of the central core (red) bounded by two circles one part of the upper 50 K domain (yellow) the other part of the relay/converter domain (blue). Each ring has a segment missing to represent the entrance to the nucleotide pocket. A·M at the top left represents the rigor actin-myosin complex with the upper 50kDa domain cleft closed to allow both sides of the cleft to make contact with actin. The nucleotide pocket entrance is open with both SW1 & 2 open. Step 1 ATP binds to myosin and myosin dissociates from actin. In detail ATP binds into the nucleotide pocket and the yellow segment rotates to close SW-1 onto the ATP in doing so the major cleft opens destroying the actin binding site leading to dissociation from actin. Step 2 The recovery stroke and ATP-hydrolysis. The blue segment rotates to bring SW-2 into contact with ATP thus rotating the converter/light-chain binding domain to complete the recovery stroke or repriming of the motor while detached from actin. Only after both SW1 and SW2 are closed is ATP hydrolysed to form the stable M.ADP.Pi complex. Step 3 Actin rebinding and the power stroke. After hydrolysis the lower 50kDa part of the cleft rebinds to actin, the cleft closes and both upper and lower 50kDa domains bind actin, this involves a rotation of the yellow segment which triggers both Pi release and the rotation of the blue segment generating a force – represented as a distortion of the converter domain. The exact order of cleft closure, Pi release and power stroke remains under debate. The location of the “elasticity” within the cross bridge is not defined. Step 4 “Sliding”. Provided the force generated in step 3 is large enough the load is moved 5-10 nm by the cross bridge working-stroke. This is shown as a relative displacement of the end of the lever relative to the actin. Step 5 Opening of the nucleotide pocket. Once the strain in the converter is dissipated by the sliding movement, the blue segment rotates further to open the nucleotide pocket and allows ADP to escape in step 6. An efficient motor requires that ADP release be limited until the cross-bridge has completed its movement or working stroke. Step 4a/5a. The strain holding cross-bridge. If the force generated in step 3 is too small to move the load no sliding takes place. The further movement of the blue segment against the load is strongly inhibited (~5-100 fold for different myosins). The degree to which this rotation is inhibited in a load bearing myosin defines the load sensitivity of the myosin. This is a function of the size of the rotation, the length of the lever arm and the stiffness of the “spring” in the structure.

Figure 2. Four groups of myosin function: fast movers, slow/efficient movers, strain sensors and gated/processive movers

For each group the correlation with the three kinetic signatures (duty ratio, K_AD/K_D and load-dependence) is indicated, together with the isomerisation constant K_CO. K_CO is the equilibrium constant for the step that opens the nucleotide binding pocket before ADP can be released. Fast movers, such as fast muscle myosin-II, have a low duty ratio and load-dependence and a high thermodynamic coupling ratio (K_AD/K_D) and K_CO value. Gated myosins, such as myosin 5a, have a high duty ratio and load-dependence but low values for the thermodynamic coupling and K_CO. The diagram is drawn to emphasise the graded nature of the change in the parameters across the series. There is not a step change in the values and therefore the boundary between each type of motor function is not exact.