news & views MOLECULAR MOTORS
Myosins move ahead of the pack
An artificial motor protein with loosely coordinated subunits can travel at high speed and over long distances.
David S. Tsao and Michael R. Diehl
T
he internal organization of cells is regulated by a variety of different mechanochemical enzymes called motor proteins. These nanomechanical machines function by consuming ATP as fuel to shuttle vesicles, organelles and other important subcellular materials directionally along cytoskeletal filament tracks1. The transport of these commodities can be quite efficient because many motor proteins are capable of moving at high velocities (~1 μm s–1) and can transport their cargoes over appreciable distances (~1 μm) compared with the dimensions of the cell. This impressive performance is often attributed to a tight mechanochemical coupling between the catalytic/motor domains within a motor molecule, which serves to coordinate the mechanochemical cycles of each domain as a motor steps along its filament tracks (Fig. 1a)2,3. Yet, writing in Nature Nanotechnology, Zev Bryant and co-workers from Stanford University4 now show that by engineering motor proteins to have loose coupling between motor domains, their speed and long-distance capabilities can be enhanced significantly. The researchers were able to create high-performance motors by modifying naturally occurring dimeric myosinVI and myosinXI proteins that, like many biological motors, possess two motor domains (heads) connected by a coiled-coiled stalk. The new myosin constructs incorporate two additional heads to enhance their binding affinity towards their filament tracks. The four heads were also linked together via flexible polypeptide elements to increase their diffusive freedom and to allow each head to bind the filament freely. These adaptations allowed the engineered myosin constructs to travel distances of several micrometres at velocities of up to 10 μm s–1. Such remarkable performance shows that maintaining tight mechanical coordination between motor heads is not essential for creating motors that are both fast and highly processive (that is, capable of completing multiple mechanochemical cycles/steps while remaining filament-bound). Similar effects had previously been found with two- and three-headed kinesins complexes5. In those cases, the coupling of individual
a
b
Pi
ATP
ADP 1×
2×
10×
Figure 1 | Strategies to enhance the performance of molecular motors. a, Tight coupling between the motor/head domains (red and yellow circles) where ATP hydrolysis is catalysed in many biological motors, like kinesin-1, helps coordinate the motions of each head. This communication between the motor heads is widely believed to facilitate fast, directional and processive stepping along cytoskeletal filament tracks. Run lengths and velocities under applied loads can be further enhanced when multiple motors are coupled to a common cargo or connected via a molecular scaffold (blue coil). Pi, phosphate ion. b, The gains described in a are much smaller than those achieved by integrating four myosinVI or myosinXI heads (red spheres) in a single multivalent motor complex if the linkages between these domains are suitably compliant.
kinesin motor domains through long (~125 amino acids) and highly flexible elastic linkages was found to accelerate ATP hydrolysis as well as the rate-limiting step that determines the motor velocity, resulting in a twofold increase in the gliding velocities along the filament (Fig. 1b). However, the velocities were still quite low. In contrast, the myosin complexes generated by Bryant and colleagues are among the fastest processive motors known so far, and can travel up to 10 times farther than their two-headed, wildtype counterparts. In previous studies, four-headed motor systems have also been created by connecting two separate dimeric motors together using protein- and DNA-based molecular scaffolds (Fig. 1b)6–9. These multiprotein complexes were developed to characterize mechanisms governing the motions of vesicles and organelles inside cells, which are often transported by motor teams
NATURE NANOTECHNOLOGY | VOL 9 | JANUARY 2014 | www.nature.com/naturenanotechnology
© 2014 Macmillan Publishers Limited. All rights reserved
containing multiple copies of the same or even different types of motor 10. Velocity and force production enhancements in these multi-motor systems are often quite small and strongly dependent on the type of motor 6,11. Although there is some evidence that the individual motors within these complexes can function synergistically when the motors transport cargoes against large applied loads6, their velocity is typically of the same magnitude as that of single-motor molecules at low applied loads11,12, whereas run lengths are, at most, twice as large. The striking performances of the Stanford Group’s multi-head myosins suggest that controlling the local coupling of individual catalytic domains may be a more effective way to custom-tailor the properties of motor systems than assembling complete motors into teams. It is important to note, however, that these myosins were tested in the absence of a load. Therefore, it will be interesting to 9
news & views see how different designs influence the ability to generate a force against an external load. Finally, the use of natural and synthetic motors as actuators and transport machinery in nanomechanical devices has also been explored. The modular approach adopted by Bryant and colleagues provides a general framework for tuning the properties of protein-based motors for these applications. The same principles can potentially be applied to configure the properties of unnatural motors built from synthetic oligonucleotides or organic molecules. Although these technologies are likely to
take time to fully mature, the ability to circumvent natural design principles should open important avenues to both customize and even adapt the properties of motors for realistic applications. ❐ David S. Tsao and Michael R. Diehl are at the Department of Bioengineering and Chemistry of Rice University, Houston, Texas 77030, USA. e-mail: [email protected] References 1. Vale, R. D. Cell 112, 467–480 (2003). 2. Purcell, T. J., Sweeney, H. L. & Spudich J. A. Proc. Natl Acad. Sci. USA 102, 13873–13878 (2005).
3. Toprak, E., Yildiz, A., Hoffman, M. T., Rosenfeldd, S. S. & Selvin, P. R. Proc. Natl Acad. Sci. USA 106, 12717–12722 (2009). 4. Schindler, T. D., Chen, L., Lebel, P., Nakamura, M. & Bryant, Z. Nature Nanotech. 9, 33–38 (2014). 5. Diehl, M. R., Zhang, K., Lee, H. J. & Tirrell, D. A. Science 311, 1468–1471 (2006). 6. Jamison, D. K., Driver, J. W. & Diehl, M. R. J. Biol. Chem. 287, 3357–3365 (2012). 7. Furuta, K. et al. Proc. Natl Acad. Sci. USA 110, 501–506 (2013). 8. Derr, N. D. et al. Science 338, 662–665 (2012). 9. Beeg, J., Klumpp, S., Dimova, R., Gracia, R. S., Unger, E. & Lipowsky, R. Biophys. J. 94, 532–541 (2008). 10. Barlan, K., Rossow, M. J. & Gelfand, V. I. Curr. Opin. Cell Biol. 25, 1–6 (2013). 11. Lu, H. et al. J. Biol. Chem. 287, 27753–27761 (2012). 12. Rogers, A. R., Driver, J. W., Constantinou, P. E., Jamison, D. K. & Diehl, M. R. Phys. Chem. Chem. Phys. 11, 4882–4889 (2009).
MOLECULAR MOTORS
On track with nanotubes
DNA motors can transport CdS nanoparticles along tracks made of carbon nanotubes.
Anand Jagota
A
goal of nanotechnology is to build molecular-scale machines that can run without intervention. This is a problem that evolution has faced, and solved with, for example, the systems that operate in cells. A living cell is a bit like a medieval walled city, with its enclosing membrane the ramparts. The ‘city’ inside these ‘walls’ bustles with life and commerce, and ‘goods’ have to be transported from point to point within it. The solution to this is a network of ‘roads’, or one-dimensional tracks, and vehicles that run on these tracks carrying cargo1. Inside mammalian cells, the tracks are usually long filamentous proteins — actin or microtubules — and the vehicles are motor proteins such as kinesin and dynein2, which are fuelled by adenosine triphosphate (ATP) molecules. Often the motors can move in both directions on the track and overall steady-state motion a
b
in one direction results from a difference in relative time spent going in one direction versus the other. Therefore, the motors use some form of biased, random (or Brownian) motion, and the tracks themselves usually have a ‘polarity’ to them, which directs the motion. Although natural molecular motors are typically made of proteins, to try and recreate such capabilities in the laboratory researchers have often turned to DNA. This is because the robust Watson–Crick base-pairing rules of DNA allows synthetic motors to be developed with programmable control. This allows complex motions to be created and even synthesis steps to be built in3–5. These molecular motor systems have three essential elements: the motor, the track on which the motor moves, and a source of energy, all of which are typically made from DNA or RNA. Writing in c
d
Nature Nanotechnology, Jong Hyun Choi and colleagues at Purdue University have now developed a DNA motor that runs autonomously on tracks of carbon nanotubes decorated with RNA molecules6. Over the past decade, a variety of DNA-based motors that run on DNA-based tracks have been demonstrated. As well as their programmability, these motors are different from the protein-based cellular motors in a number of ways. For example, in most cases the motors are not driven by ATP hydrolysis but by some transformation of DNA itself. Some motors use an enzyme that cuts DNA to move DNA cargo along a DNA track7. A related design uses a DNA enzyme to cleave an RNA molecule8. Because the track usually has no polarity or orientation, something needs to be done to break the symmetry so as to prescribe a direction of motion. Often the solution is e
Figure 1 | DNA motors on carbon nanotube tracks. a, The DNA enzyme motors, which consist of a catalytic core (green) and two recognition arms (red), are attached to nanoparticle cargoes (yellow) and can bind to RNA molecules (blue) that decorate the carbon nanotube (black) tracks. b, Once bound, the DNA enzyme can cut the RNA molecules in two. c,d, The shorter of these two RNA strands then dissociates and diffuses away (c), and the unpaired DNA arm binds to the next RNA molecule on the nanotube (d). e, The longer DNA stand then gets dragged over to the new RNA strand, and the motor is ready to take another step. 10
NATURE NANOTECHNOLOGY | VOL 9 | JANUARY 2014 | www.nature.com/naturenanotechnology
© 2014 Macmillan Publishers Limited. All rights reserved
Myosins move ahead of the pack
An artificial motor protein with loosely coordinated subunits can travel at high speed and over long distances.
David S. Tsao and Michael R. Diehl
T
he internal organization of cells is regulated by a variety of different mechanochemical enzymes called motor proteins. These nanomechanical machines function by consuming ATP as fuel to shuttle vesicles, organelles and other important subcellular materials directionally along cytoskeletal filament tracks1. The transport of these commodities can be quite efficient because many motor proteins are capable of moving at high velocities (~1 μm s–1) and can transport their cargoes over appreciable distances (~1 μm) compared with the dimensions of the cell. This impressive performance is often attributed to a tight mechanochemical coupling between the catalytic/motor domains within a motor molecule, which serves to coordinate the mechanochemical cycles of each domain as a motor steps along its filament tracks (Fig. 1a)2,3. Yet, writing in Nature Nanotechnology, Zev Bryant and co-workers from Stanford University4 now show that by engineering motor proteins to have loose coupling between motor domains, their speed and long-distance capabilities can be enhanced significantly. The researchers were able to create high-performance motors by modifying naturally occurring dimeric myosinVI and myosinXI proteins that, like many biological motors, possess two motor domains (heads) connected by a coiled-coiled stalk. The new myosin constructs incorporate two additional heads to enhance their binding affinity towards their filament tracks. The four heads were also linked together via flexible polypeptide elements to increase their diffusive freedom and to allow each head to bind the filament freely. These adaptations allowed the engineered myosin constructs to travel distances of several micrometres at velocities of up to 10 μm s–1. Such remarkable performance shows that maintaining tight mechanical coordination between motor heads is not essential for creating motors that are both fast and highly processive (that is, capable of completing multiple mechanochemical cycles/steps while remaining filament-bound). Similar effects had previously been found with two- and three-headed kinesins complexes5. In those cases, the coupling of individual
a
b
Pi
ATP
ADP 1×
2×
10×
Figure 1 | Strategies to enhance the performance of molecular motors. a, Tight coupling between the motor/head domains (red and yellow circles) where ATP hydrolysis is catalysed in many biological motors, like kinesin-1, helps coordinate the motions of each head. This communication between the motor heads is widely believed to facilitate fast, directional and processive stepping along cytoskeletal filament tracks. Run lengths and velocities under applied loads can be further enhanced when multiple motors are coupled to a common cargo or connected via a molecular scaffold (blue coil). Pi, phosphate ion. b, The gains described in a are much smaller than those achieved by integrating four myosinVI or myosinXI heads (red spheres) in a single multivalent motor complex if the linkages between these domains are suitably compliant.
kinesin motor domains through long (~125 amino acids) and highly flexible elastic linkages was found to accelerate ATP hydrolysis as well as the rate-limiting step that determines the motor velocity, resulting in a twofold increase in the gliding velocities along the filament (Fig. 1b). However, the velocities were still quite low. In contrast, the myosin complexes generated by Bryant and colleagues are among the fastest processive motors known so far, and can travel up to 10 times farther than their two-headed, wildtype counterparts. In previous studies, four-headed motor systems have also been created by connecting two separate dimeric motors together using protein- and DNA-based molecular scaffolds (Fig. 1b)6–9. These multiprotein complexes were developed to characterize mechanisms governing the motions of vesicles and organelles inside cells, which are often transported by motor teams
NATURE NANOTECHNOLOGY | VOL 9 | JANUARY 2014 | www.nature.com/naturenanotechnology
© 2014 Macmillan Publishers Limited. All rights reserved
containing multiple copies of the same or even different types of motor 10. Velocity and force production enhancements in these multi-motor systems are often quite small and strongly dependent on the type of motor 6,11. Although there is some evidence that the individual motors within these complexes can function synergistically when the motors transport cargoes against large applied loads6, their velocity is typically of the same magnitude as that of single-motor molecules at low applied loads11,12, whereas run lengths are, at most, twice as large. The striking performances of the Stanford Group’s multi-head myosins suggest that controlling the local coupling of individual catalytic domains may be a more effective way to custom-tailor the properties of motor systems than assembling complete motors into teams. It is important to note, however, that these myosins were tested in the absence of a load. Therefore, it will be interesting to 9
news & views see how different designs influence the ability to generate a force against an external load. Finally, the use of natural and synthetic motors as actuators and transport machinery in nanomechanical devices has also been explored. The modular approach adopted by Bryant and colleagues provides a general framework for tuning the properties of protein-based motors for these applications. The same principles can potentially be applied to configure the properties of unnatural motors built from synthetic oligonucleotides or organic molecules. Although these technologies are likely to
take time to fully mature, the ability to circumvent natural design principles should open important avenues to both customize and even adapt the properties of motors for realistic applications. ❐ David S. Tsao and Michael R. Diehl are at the Department of Bioengineering and Chemistry of Rice University, Houston, Texas 77030, USA. e-mail: [email protected] References 1. Vale, R. D. Cell 112, 467–480 (2003). 2. Purcell, T. J., Sweeney, H. L. & Spudich J. A. Proc. Natl Acad. Sci. USA 102, 13873–13878 (2005).
3. Toprak, E., Yildiz, A., Hoffman, M. T., Rosenfeldd, S. S. & Selvin, P. R. Proc. Natl Acad. Sci. USA 106, 12717–12722 (2009). 4. Schindler, T. D., Chen, L., Lebel, P., Nakamura, M. & Bryant, Z. Nature Nanotech. 9, 33–38 (2014). 5. Diehl, M. R., Zhang, K., Lee, H. J. & Tirrell, D. A. Science 311, 1468–1471 (2006). 6. Jamison, D. K., Driver, J. W. & Diehl, M. R. J. Biol. Chem. 287, 3357–3365 (2012). 7. Furuta, K. et al. Proc. Natl Acad. Sci. USA 110, 501–506 (2013). 8. Derr, N. D. et al. Science 338, 662–665 (2012). 9. Beeg, J., Klumpp, S., Dimova, R., Gracia, R. S., Unger, E. & Lipowsky, R. Biophys. J. 94, 532–541 (2008). 10. Barlan, K., Rossow, M. J. & Gelfand, V. I. Curr. Opin. Cell Biol. 25, 1–6 (2013). 11. Lu, H. et al. J. Biol. Chem. 287, 27753–27761 (2012). 12. Rogers, A. R., Driver, J. W., Constantinou, P. E., Jamison, D. K. & Diehl, M. R. Phys. Chem. Chem. Phys. 11, 4882–4889 (2009).
MOLECULAR MOTORS
On track with nanotubes
DNA motors can transport CdS nanoparticles along tracks made of carbon nanotubes.
Anand Jagota
A
goal of nanotechnology is to build molecular-scale machines that can run without intervention. This is a problem that evolution has faced, and solved with, for example, the systems that operate in cells. A living cell is a bit like a medieval walled city, with its enclosing membrane the ramparts. The ‘city’ inside these ‘walls’ bustles with life and commerce, and ‘goods’ have to be transported from point to point within it. The solution to this is a network of ‘roads’, or one-dimensional tracks, and vehicles that run on these tracks carrying cargo1. Inside mammalian cells, the tracks are usually long filamentous proteins — actin or microtubules — and the vehicles are motor proteins such as kinesin and dynein2, which are fuelled by adenosine triphosphate (ATP) molecules. Often the motors can move in both directions on the track and overall steady-state motion a
b
in one direction results from a difference in relative time spent going in one direction versus the other. Therefore, the motors use some form of biased, random (or Brownian) motion, and the tracks themselves usually have a ‘polarity’ to them, which directs the motion. Although natural molecular motors are typically made of proteins, to try and recreate such capabilities in the laboratory researchers have often turned to DNA. This is because the robust Watson–Crick base-pairing rules of DNA allows synthetic motors to be developed with programmable control. This allows complex motions to be created and even synthesis steps to be built in3–5. These molecular motor systems have three essential elements: the motor, the track on which the motor moves, and a source of energy, all of which are typically made from DNA or RNA. Writing in c
d
Nature Nanotechnology, Jong Hyun Choi and colleagues at Purdue University have now developed a DNA motor that runs autonomously on tracks of carbon nanotubes decorated with RNA molecules6. Over the past decade, a variety of DNA-based motors that run on DNA-based tracks have been demonstrated. As well as their programmability, these motors are different from the protein-based cellular motors in a number of ways. For example, in most cases the motors are not driven by ATP hydrolysis but by some transformation of DNA itself. Some motors use an enzyme that cuts DNA to move DNA cargo along a DNA track7. A related design uses a DNA enzyme to cleave an RNA molecule8. Because the track usually has no polarity or orientation, something needs to be done to break the symmetry so as to prescribe a direction of motion. Often the solution is e
Figure 1 | DNA motors on carbon nanotube tracks. a, The DNA enzyme motors, which consist of a catalytic core (green) and two recognition arms (red), are attached to nanoparticle cargoes (yellow) and can bind to RNA molecules (blue) that decorate the carbon nanotube (black) tracks. b, Once bound, the DNA enzyme can cut the RNA molecules in two. c,d, The shorter of these two RNA strands then dissociates and diffuses away (c), and the unpaired DNA arm binds to the next RNA molecule on the nanotube (d). e, The longer DNA stand then gets dragged over to the new RNA strand, and the motor is ready to take another step. 10
NATURE NANOTECHNOLOGY | VOL 9 | JANUARY 2014 | www.nature.com/naturenanotechnology
© 2014 Macmillan Publishers Limited. All rights reserved
