Structural dynamics of myosin 5 during processive motion revealed by interferometric scattering microscopy

Andrecka, Joanna; Arroyo, Jaime Ortega; Takagi, Yasuharu; Wit, Gabrielle de; Fineberg, Adam; MacKinnon, Lachlan; Young, Gavin; Sellers, James R.; Kukura, Philipp

doi:10.7554/elife.05413

Cited by 73 publications

(88 citation statements)

References 55 publications

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“…The free rotation of the TL is in accordance with the motion observed in many single-molecule experiments on myosin-V (35,36). Some of the trajectories have shown recursive motions of the TL, where the leg, instead of binding to the next actin-binding site, goes back to the previous site.…”

Section: Resultssupporting

confidence: 62%

“…Application of higher loads increases both the diffusive motion and the bound dwell time for the myosin-V legs. These results reveal that our model of myosin with an energydemanding PS is actually capable of providing qualitatively similar walking motions as observed in experiments for both unloaded and loaded cases (35,36). However, it should be noted that the time for a single myosin-V step is much faster in our simulations (especially the phase where both ADP-bound legs are bound to actin) compared with experimental observations.…”

Section: Dynamics Of Myosin-v Under Constant Load: Dependence Of Stallcontrasting

confidence: 45%

“…We also observe substep features (most evident in single-leg trajectories of Fig. 3 B and C), as found in single-molecule and AFM experiments (35,36). The motion of the TL follows a substep of ∼40-50 nm, most of which is diffusive in nature after it releases the actin-binding site.…”

Section: Resultsmentioning

confidence: 48%

“…The Coupling of the P i Release and the Lever Arm Movement of Myosin-V. One of our aims has been to explore whether having an energy-demanding PS interweaved with the chemical steps of ATP binding, P i , and ADP release and actin-binding/release events can produce the myosin-V motion toward the plus end, as has been observed in numerous experiments (35)(36)(37). This issue was explored by running LD simulations under different conditions.…”

Section: Resultsmentioning

confidence: 99%

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Simulating the dynamics of the mechanochemical cycle of myosin-V

Mukherjee

Alhadeff

Warshel

2017

Proc. Natl. Acad. Sci. U.S.A.

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The detailed dynamics of the cycle of myosin-V are explored by simulation approaches, examining the nature of the energy-driven motion. Our study started with Langevin dynamics (LD) simulations on a very coarse landscape with a single rate-limiting barrier and reproduced the stall force and the hand-over-hand dynamics. We then considered a more realistic landscape and used timedependent Monte Carlo (MC) simulations that allowed trajectories long enough to reproduce the force/velocity characteristic sigmoidal correlation, while also reproducing the hand-over-hand motion. Overall, our study indicated that the notion of a downhill lever-up to lever-down process (popularly known as the powerstroke mechanism) is the result of the energetics of the complete myosin-V cycle and is not the source of directional motion or force generation on its own. The present work further emphasizes the need to use well-defined energy landscapes in studying molecular motors in general and myosin in particular.M yosin constitutes a superfamily of molecular motors comprising both the nonprocessive single-headed motors that are efficient in generating force on the actin filaments (e.g., myosin-II) and highly processive double-headed motors that can transport cellular load using tracks laid out by the cytoskeletal actin filaments (e.g., myosin-V, myosin-VI) (1, 2). The generation of force and unidirectional motion in each of the myosin molecules predominantly comprises tightly coupled events involving (i) binding and hydrolysis of ATP, followed by release of the products ADP and inorganic phosphate (P i ), occurring at the nucleotide-binding domain; (ii) binding and release of actin through the actin-binding domain; and (iii) a large conformational change of the lever arm that generates mechanical motion. Although the basic mechanochemical cycle is conserved in all members of the myosin superfamily, the exact nature of the coupling between the above steps in myosins, which decides the force-generating and load-bearing characteristics of different members, is unknown (SI Background).Numerous experiments, including kinetics and thermodynamic studies (3), high-resolution structural studies (4), electron microscopy studies (5), atomic force microscopy (AFM), and singlemolecule experiments (2, 6, 7), have advanced our understanding of the action of the system. Although details about the dynamical nature of the myosin-V, along with a quantitative knowledge of the force-response, have been learned from single-molecule studies, the structural studies have provided us with an atomistic knowledge of the key conformational states involved during the cycle, namely, the lever-up (pre) and lever-down (post) states (Fig. S1). In addition, crucial information has been gained on the kinetics of the chemical and ligand-binding/release steps that make up the complete cycle. Based on this experimental information, several theoretical modeling studies (8-17) explored the mechanochemical cycle, mostly using phenomenological modeling approaches, accompanied by s...

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Section: Resultssupporting

confidence: 62%

Section: Dynamics Of Myosin-v Under Constant Load: Dependence Of Stallcontrasting

confidence: 45%

Section: Resultsmentioning

confidence: 48%

Section: Resultsmentioning

confidence: 99%

See 2 more Smart Citations

Simulating the dynamics of the mechanochemical cycle of myosin-V

Mukherjee

Alhadeff

Warshel

2017

Proc. Natl. Acad. Sci. U.S.A.

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“…Data were processed and analyzed using National Instruments LabVIEW 2011 and the FIJI distribution of ImageJ. To correct for illumination inhomogeneity and fixed pattern noise, a flat-field image was taken by running a temporal median filter over a sequence of images acquired when the sample was displaced (22). Differential imaging was achieved by subtracting sets of images temporally offset by a time Δt.…”

Section: Methodsmentioning

confidence: 99%

Visualization of the spontaneous emergence of a complex, dynamic, and autocatalytic system

Arroyo

Bissette

Kukura

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

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Autocatalytic chemical reactions are widely studied as models of biological processes and to better understand the origins of life on Earth. Minimal self-reproducing amphiphiles have been developed in this context and as an approach to de novo "bottom-up" synthetic protocells. How chemicals come together to produce living systems, however, remains poorly understood, despite much experimentation and speculation. Here, we use ultrasensitive label-free optical microscopy to visualize the spontaneous emergence of an autocatalytic system from an aqueous mixture of two chemicals. Quantitative, in situ nanoscale imaging reveals heterogeneous self-reproducing aggregates and enables the real-time visualization of the synthesis of new aggregates at the reactive interface. The aggregates and reactivity patterns observed vary together with differences in the respective environment. This work demonstrates how imaging of chemistry at the nanoscale can provide direct insight into the dynamic evolution of nonequilibrium systems across molecular to microscopic length scales.autocatalysis | label-free microscopy | interferometric scattering | protocells | emergence

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Fast Leaps between Millisecond Confinements Govern Ase1 Diffusion along Microtubules

et al. 2021

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Diffusion is the most fundamental mode of protein translocation within cells. Confined diffusion of proteins along the electrostatic potential constituted by the surface of microtubules, although modeled meticulously in molecular dynamics simulations, has not been experimentally observed in real‐time. Here, interferometric scattering microscopy is used to directly visualize the movement of the microtubule‐associated protein Ase1 along the microtubule surface at nanometer and microsecond resolution. Millisecond confinements of Ase1 and fast leaps between these positions of dwelling preferentially occurring along the microtubule protofilaments are resolved, revealing Ase1's mode of diffusive translocation along the microtubule's periodic surface. The derived interaction potential closely matches the tubulin‐dimer periodicity and the distribution of the electrostatic potential on the microtubule lattice. It is anticipated that mapping the interaction landscapes for different proteins on microtubules, finding plausible energetic barriers of different positioning and heights, can provide valuable insights into regulating the dynamics of essential cytoskeletal processes, such as intracellular cargo trafficking, cell division, and morphogenesis, all of which rely on diffusive translocation of proteins along microtubules.

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Structural dynamics of myosin 5 during processive motion revealed by interferometric scattering microscopy

Cited by 73 publications

References 55 publications

Simulating the dynamics of the mechanochemical cycle of myosin-V

Simulating the dynamics of the mechanochemical cycle of myosin-V

Visualization of the spontaneous emergence of a complex, dynamic, and autocatalytic system

Fast Leaps between Millisecond Confinements Govern Ase1 Diffusion along Microtubules

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