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Kinesin: Switch I & II and the Motor Mechanism

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Kinesin: Switch I & II and the Motor Mechanism Commentary 15 Kinesin: switch I & II and the motor mechanism F. Jon Kull1,* and Sharyn A. Endow2,‡ 1Department of Biophysics, Max-Planck Institute for Medical Research, Jahnstrasse 29, D-69120 Heidelberg Germany 2Department of Cell Biology, Duke University Medical Center, Durham, NC 27710 USA *Present address: Department of Chemistry, Dartmouth College, Hanover, NH 03755 USA ‡Author for correspondence (e-mail: [email protected]) Journal of Cell Science 115, 15-23 (2002) © The Company of Biologists Ltd Summary New crystal structures of the kinesin motors differ from transitions that occur in the kinesin motors during the ATP previously described motor-ADP atomic models, showing hydrolysis cycle. The movements of switch I residues striking changes both in the switch I region near the disrupt the water-mediated coordination of the bound nucleotide-binding cleft and in the switch II or ‘relay’ helix Mg2+, which could result in loss of Mg2+ and ADP, raising at the filament-binding face of the motor. The switch I the intriguing possibility that disruption of the switch I region, present as a short helix flanked by two loops in region leads to release of nucleotide by the kinesins. None previous motor-ADP structures, rearranges into a pseudo- of the new structures is a true motor-ATP state, however, β-hairpin or is completely disordered with melted helices probably because conformational changes at the active site to either side of the disordered switch I loop. The relay helix of the kinesins require interactions with microtubules to undergoes a rotational movement coupled to a translation stabilize the movements. that differs from the piston-like movement of the relay helix observed in myosin. The changes observed in the crystal Key words: Molecular motor, Structure/function, Conformational structures are interpreted to represent structural changes, Kinesin, Myosin Introduction more steps of ATP binding or hydrolysis induces small Life inside the cell is an incessant frenzy of rapid back-and- conformational changes in the protein that, under load, create forth movements of myriad cellular components, including strain (Howard, 2001). The strain is relieved by further changes proteins, vesicles and organelles, driven by collisions of water in the motor that produce force and then amplify the force, and other molecules (see Howard, 2001). This Brownian resulting in movement of the motor along its filament. The motion allows molecules such as newly synthesized RNAs to structural elements that undergo strain are likely to have diffuse randomly from their site of entry into the cytoplasm but spring-like or elastic properties that allow them to extend or is far too slow to move molecules the relatively large distances rotate, and then recoil back into their original conformation required in eukaryotic cells and cannot localize them to specific (Howard, 2001). Movements of the motor catalytic core are regions of the cell (Wilkie and Davis, 2001). Localization to a further expected to involve the highly conserved switch regions site within the cell requires facilitated movement, usually aided (Kull et al., 1996; Sablin et al., 1996), switch I and switch II, by one of a battery of cytoskeletal motor proteins - the myosins, so-named because of their structural homology to regions of G kinesins or dyneins. proteins that move upon nucleotide hydrolysis and exchange (Sprang, 1997). Thus, an understanding of the motor mechanism is likely to come only after workers have identified Cytoskeletal motors the spring-like or elastic elements within the motor, together Cytoskeletal motor proteins are specially built to perform their with the force-producing structural changes in the motor and transport function in the cell: one region of the protein binds the steps in the hydrolysis cycle at which they occur. to an actin filament or microtubule, hydrolyzes ATP and moves The two best-studied cytoskeletal motors, myosin and along the filament, while another region attaches to cargo kinesin, are dimeric proteins that have two catalytic domains for transport to a specific site within the cell. Cytoskeletal joined by a coiled-coil rod or stalk. These two motor proteins motors are thus specialized ATPases that transport cargo by and other highly related proteins in their respective families coordinating the hydrolysis of ATP with binding to and contain a central core of structural elements that are remarkably movement along a filament. Remarkably, these molecular similar to one another (Kull et al., 1996) (Fig. 1). Despite this motors convert the chemical energy from ATP hydrolysis structural homology, however, there are indications that the directly into mechanical energy. How do motors capture the kinesin motors differ substantially from the myosins in their energy released by nucleotide hydrolysis and turn it into mechanism of function. A fundamental difference is the work? Motor proteins are apparently capable of sensing and nucleotide-dependent interactions of the motors with their responding to the presence or absence of a γ-phosphate, and filament: myosin bound to ATP is weakly bound to or detached then transmitting this information along a pathway of from actin, whereas kinesin-ATP is strongly bound to increasingly larger conformational changes that culminates in microtubules. Conversely, myosin-ADP is strongly bound to a force-generating event. A prevailing idea is that one or actin, whereas kinesin-ADP is weakly bound to or detached 16 Journal of Cell Science 115 (1) from microtubules. A further basic difference between the motors have been elusive. Almost all of the crystal structures kinesins and myosins is that myosin hydrolyzes ATP while solved so far are complexed to ADP and are interpreted to detached from actin, whereas kinesin hydrolyses ATP while represent weak-binding or detached ADP states (reviewed by attached to the microtubule. But, for both motors, the rate- Sack et al., 1999). The kinesin-ADP models do differ from one limiting step in the hydrolysis cycle is accelerated by binding another, indicating that they may represent structural of the motor to its filament, which results in a characteristic transitions within the ADP state, which could help identify the actin- or microtubule-activated ATPase activity that underlies mechanical elements of the motor that undergo conformational the ability of the motor to move along its filament. changes during the hydrolysis cycle. For example, human kinesin (Kull et al., 1996) and rat kinesin (Kozielski et al., Myosin Work on myosin has identified three distinct conformations of the motor that are thought to correspond to different nucleotide states (Rayment et al., 1993a; Fisher et al., 1995; Dominguez et al., 1998; Houdusse et al., 1999; Houdusse et al., 2000). These have been classified as detached, near rigor, and transition forms (Houdusse et al., 2000) (Fig. 2), all of which bind weakly to actin. The nucleotide bound to the motor is not always an indication of the motor state, since the detached form can be crystallized with either ADP or ATP bound to the active site (Houdusse et al., 1999; Houdusse et al., 2000). The three forms show major differences in the position of the rod-like lever arm, as well as a striking change in the ‘converter’ region at the base of the lever arm. The converter, a rigid α/β subdomain of 67 residues that includes the first three turns of the helical rod (Fig. 1), is thought to convert movements at the nucleotide-binding cleft of the motor, which are transmitted by the adjacent switch II or ‘relay’ helix and the SH1 helix, into the swinging of the lever arm. The SH1 helix, which is also adjacent to the converter, is intact in the near-rigor and transition structures but disordered in the detached form, which has been interpreted to be a motor-ATP state (Houdusse et al., 1999). This mobility of the SH1 helix reflects its role in transmitting and directing movements of the relay helix to the converter domain and lever arm. Structural changes at the nucleotide-binding cleft of myosin are thus coupled to movements of the relay helix, SH1 helix, converter domain and the lever arm (Fig.1). The power stroke is thought to correspond to the swinging of the lever arm (Rayment et al., 1993a; Rayment et al., 1993b); however, the structural elements that undergo strain and the conformational changes in myosin that produce force have not yet been identified. Kinesin motors By comparison with myosin, the structural states of the kinesin Fig. 1. Structural comparison of kinesin and myosin. (A) Front view of kinesin (rat monomeric KHC, PDB 2KIN) (Sack et al., 1997) and myosin II (scallop S1-ADP·VO4, PDB 1DFL) (Houdusse et al., 2000). Core β-strands with adjacent nucleotide-binding (P-loop) or switch regions are colored green (P-loop), purple (switch I) and cyan (switch II). The helix following switch II, the ‘relay’ helix, is red in the two motors. Other β-strands and α-helices common to kinesin and myosin are shown in dark blue-gray and unique areas are in light gray. The myosin converter and kinesin neck helix and neck linker are shown in pink-purple. The myosin lever arm is tan. (B) Back view, rotated 180° from the view in A, showing the filament-binding face of the motors. Switch II at the active site of myosin is connected to the relay helix, which interacts with the converter and lever arm at its other end - the converter can therefore convert changes at the active site into movements of the lever arm. Kinesin: switch I & II and the motor mechanism 17 1997) show little change in the nucleotide-binding site (Fig. myosin) forms a hydrogen bond with an oxygen from the γ- 3A), but helices α4 and α5 are displaced in rat kinesin relative phosphate; (2) a conserved serine (or threonine) residue from to human kinesin (Fig.
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