Direct observation of the myosin Va recovery stroke that contributes to unidirectional stepping along actin.
Myosins are ATP-driven linear molecular motors that work as cellular force generators, transporters, and force sensors. These functions are driven by large-scale nucleotide-dependent conformational changes, termed "strokes"; the "power stroke" is the force-generating swinging of...
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Public Library of Science (PLoS)
2011-04-01
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| Series: | PLoS Biology |
| Online Access: | https://journals.plos.org/plosbiology/article/file?id=10.1371/journal.pbio.1001031&type=printable |
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| author | Katsuyuki Shiroguchi Harvey F Chin Diane E Hannemann Eiro Muneyuki Enrique M De La Cruz Kazuhiko Kinosita |
| author_facet | Katsuyuki Shiroguchi Harvey F Chin Diane E Hannemann Eiro Muneyuki Enrique M De La Cruz Kazuhiko Kinosita |
| author_sort | Katsuyuki Shiroguchi |
| collection | DOAJ |
| description | Myosins are ATP-driven linear molecular motors that work as cellular force generators, transporters, and force sensors. These functions are driven by large-scale nucleotide-dependent conformational changes, termed "strokes"; the "power stroke" is the force-generating swinging of the myosin light chain-binding "neck" domain relative to the motor domain "head" while bound to actin; the "recovery stroke" is the necessary initial motion that primes, or "cocks," myosin while detached from actin. Myosin Va is a processive dimer that steps unidirectionally along actin following a "hand over hand" mechanism in which the trailing head detaches and steps forward ∼72 nm. Despite large rotational Brownian motion of the detached head about a free joint adjoining the two necks, unidirectional stepping is achieved, in part by the power stroke of the attached head that moves the joint forward. However, the power stroke alone cannot fully account for preferential forward site binding since the orientation and angle stability of the detached head, which is determined by the properties of the recovery stroke, dictate actin binding site accessibility. Here, we directly observe the recovery stroke dynamics and fluctuations of myosin Va using a novel, transient caged ATP-controlling system that maintains constant ATP levels through stepwise UV-pulse sequences of varying intensity. We immobilized the neck of monomeric myosin Va on a surface and observed real time motions of bead(s) attached site-specifically to the head. ATP induces a transient swing of the neck to the post-recovery stroke conformation, where it remains for ∼40 s, until ATP hydrolysis products are released. Angle distributions indicate that the post-recovery stroke conformation is stabilized by ≥ 5 k(B)T of energy. The high kinetic and energetic stability of the post-recovery stroke conformation favors preferential binding of the detached head to a forward site 72 nm away. Thus, the recovery stroke contributes to unidirectional stepping of myosin Va. |
| format | Article |
| id | doaj-art-5c64e2bad38048d58846777faa7a7c31 |
| institution | DOAJ |
| issn | 1544-9173 1545-7885 |
| language | English |
| publishDate | 2011-04-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS Biology |
| spelling | doaj-art-5c64e2bad38048d58846777faa7a7c312025-08-20T03:10:02ZengPublic Library of Science (PLoS)PLoS Biology1544-91731545-78852011-04-0194e100103110.1371/journal.pbio.1001031Direct observation of the myosin Va recovery stroke that contributes to unidirectional stepping along actin.Katsuyuki ShiroguchiHarvey F ChinDiane E HannemannEiro MuneyukiEnrique M De La CruzKazuhiko KinositaMyosins are ATP-driven linear molecular motors that work as cellular force generators, transporters, and force sensors. These functions are driven by large-scale nucleotide-dependent conformational changes, termed "strokes"; the "power stroke" is the force-generating swinging of the myosin light chain-binding "neck" domain relative to the motor domain "head" while bound to actin; the "recovery stroke" is the necessary initial motion that primes, or "cocks," myosin while detached from actin. Myosin Va is a processive dimer that steps unidirectionally along actin following a "hand over hand" mechanism in which the trailing head detaches and steps forward ∼72 nm. Despite large rotational Brownian motion of the detached head about a free joint adjoining the two necks, unidirectional stepping is achieved, in part by the power stroke of the attached head that moves the joint forward. However, the power stroke alone cannot fully account for preferential forward site binding since the orientation and angle stability of the detached head, which is determined by the properties of the recovery stroke, dictate actin binding site accessibility. Here, we directly observe the recovery stroke dynamics and fluctuations of myosin Va using a novel, transient caged ATP-controlling system that maintains constant ATP levels through stepwise UV-pulse sequences of varying intensity. We immobilized the neck of monomeric myosin Va on a surface and observed real time motions of bead(s) attached site-specifically to the head. ATP induces a transient swing of the neck to the post-recovery stroke conformation, where it remains for ∼40 s, until ATP hydrolysis products are released. Angle distributions indicate that the post-recovery stroke conformation is stabilized by ≥ 5 k(B)T of energy. The high kinetic and energetic stability of the post-recovery stroke conformation favors preferential binding of the detached head to a forward site 72 nm away. Thus, the recovery stroke contributes to unidirectional stepping of myosin Va.https://journals.plos.org/plosbiology/article/file?id=10.1371/journal.pbio.1001031&type=printable |
| spellingShingle | Katsuyuki Shiroguchi Harvey F Chin Diane E Hannemann Eiro Muneyuki Enrique M De La Cruz Kazuhiko Kinosita Direct observation of the myosin Va recovery stroke that contributes to unidirectional stepping along actin. PLoS Biology |
| title | Direct observation of the myosin Va recovery stroke that contributes to unidirectional stepping along actin. |
| title_full | Direct observation of the myosin Va recovery stroke that contributes to unidirectional stepping along actin. |
| title_fullStr | Direct observation of the myosin Va recovery stroke that contributes to unidirectional stepping along actin. |
| title_full_unstemmed | Direct observation of the myosin Va recovery stroke that contributes to unidirectional stepping along actin. |
| title_short | Direct observation of the myosin Va recovery stroke that contributes to unidirectional stepping along actin. |
| title_sort | direct observation of the myosin va recovery stroke that contributes to unidirectional stepping along actin |
| url | https://journals.plos.org/plosbiology/article/file?id=10.1371/journal.pbio.1001031&type=printable |
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