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Kinesin Action Crystallized in Two Key States

Without infrastructure, business grinds to a halt, and the business of a cell is no exception. Cells require an internal transportation system reliable and flexible enough to accommodate both the routine movement of organelles and the dramatic choreography of mitosis. In response, nature has engineered an intracellular rail system of sorts, in which a "motor" enzyme called kinesin hauls chromosomes and other cellular freight along microtubule tracks. Disruption of this system can lead to certain neurological disorders as well as cancer. To better understand how this system works, a team of researchers from the University of Tokyo and the University of California, San Francisco, working at the ALS and Stanford Synchrotron Radiation Laboratory, compared the structures of the kinesin mechanism when crystallized in two functionally critical states.

dimeric kinesin movement
Movement of dimeric kinesin along a microtubule. One head (yellow) attaches to the microtubule surface (gray) while the other head (blue), attached to the neck linker (purple), swings forward.

Kinesin's kinetic energy comes from the energy released when adenosine triphosphate (ATP) is hydrolyzed to produce adenosine diphosphate (ADP). The energy is thought to drive a stepping sequence in the kinesin molecule, which, in its conventional dimeric form, has a two-headed bilateral symmetry: one "head" remains attached to the microtubule surface while the other is free to move forward. However, the core region of a monomeric form of kinesin (KIF1A) has been observed to take multiple steps before detaching, suggesting that the KIF1A core region plays a vital role in kinesin's mobility.

Biological Nanomachines

When you hear futurists speak with great excitement about the promise of nanotechnology, remember that they are speaking of artificial nanomachines, because natural nanomachines already exist in every cell of your body. Kinesin, for example, is an amazing motor enzyme that "walks" in eight-nanometer steps along microscopic filaments called microtubules, dragging along various cargoes of mitochondria, chromosomes, or other cellular organelles. Nerve cells rely on kinesin to carry neurotransmitters out to tendril-like extremities that can extend several feet from the cell's main body. When a cell is ready to divide, its tangle of microtubules align into a "spindle," and it is kinesin that propels the chromosome pairs into their separate corners. Needless to say, breakdowns in this complex transport system can have serious health consequences. By looking closely at kinesin's molecular structure in two key states, scientists hope to add to the growing store of knowledge about these nanoscale engines of life.

In this work, the researchers determined the structure of the KIF1A core region bound to ADP (to 2.2 Å) and to a nonhydrolyzable analogue of ATP (to 2.0 Å). Previously, all attempts to crystallize the ATP-like complex had failed. The research team's success in crystallizing this complex provided an excellent opportunity to compare "snapshots" of two key moments in the kinesin stepping sequence. Although the two structures look very similar overall, marked differences were observed in two "switch" regions near the ATP/ADP binding site.

Switch regions in ADP and ATP states
Comparison of KIF1A switch regions in the ADP (yellow) and ATP-like (red) states. Part of the bound ATP is shown in gray.

In the switch I region, a helix flanked by two short loops in the ADP state forms a short b-hairpin structure in the ATP-like state. The switch II region shows a series of structural changes, including the partial unwinding of a helix and its rotation by roughly 20 degrees in the ATP-like state. To visualize the effect of these changes when the kinesin is attached to a microtubule, the researchers embedded the molecular structures within cryo-electron microscope (cryo-EM) images of kinesin in contact with a microtubule surface. The results indicate that the switch II region actually remains fixed relative to the microtubule, while the rest of the KIF1A core rotates by about 20 degrees in the opposite direction. The authors suggest that this rotation binds the kinesin more tightly to the microtubule and creates a directional bias by pointing the tip of the KIF1A core in the direction of motion.

Kineison molecular model

Rotation of KIF1A core (yellow) relative to microtubule surface (gray). Left: Kinesin molecular model embedded within a cryo-EM map of kinesin. Right: Kinesin orientation in ADP (pink line) and ATP-like (red line) states.

Another important difference between the two states was found in the "neck linker" domain, which connects the two heads in the dimeric form of kinesin (and which was "grafted" onto the KIF1A monomer for this experiment). The structures show that the linker is "docked" near the core in the ATP-like state but is undocked and disordered in the ADP state. This finding supports the hypothesis that the neck linker is a crucial part of the mechanism that drives kinesin. In this view, the kinesin core is a modular base onto which different types of neck domains serve as mechanical amplifiers and transmitters (transmissions and drive shafts) whose exact function depends on the kinesin variant to which the neck linker belongs. In general, these results confirm expectations—arrived at by analogy to previously studied, structurally similar proteins—that the conformational changes observed in KIF1A are modular and extend to all kinesins. They also suggest a rationale for kinesin's tendency to move in a given direction along a microtubule.

Research conducted by M. Kikkawa, Y. Okada, H. Yajima, and N. Hirokawa (University of Tokyo); E.P. Sablin and R.J. Fletterick (University of California, San Francisco).

Research funding: Ministry of Education, Science, Sports, and Culture of Japan; National Institutes of Health. Operation of the ALS is supported by the U.S. Deptartment of Energy, Office of Basic Energy Sciences.

Publication about this research: M. Kikkawa, E.P. Sablin, Y. Okada, H. Yajima, R.J. Fletterick, N. Hirokawa, "Switch-based mechnanism of kinesin motors," Nature 411, 439 (2001).

ALSNews Vol. 198, May 8, 2002

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