Recent studies provide strong evidence that single class V myosin molecules transport vesicles and organelles processively along F-actin, taking several 36-nm steps, hand over hand, for each diffusional encounter. We demonstrated that the ATPase activity of myosin required calcium for maximal activity and showed that, in the absence of calcium, myosin V adopted a folded, inactive structure. We have used cryo-electron microscopy to examine the structure of myosin V that is walking on actin. These images give clear pictures of myosin V molecules with both heads attached to actin and will allow us to make observations about lever arm position and stiffness. We have used a fluorescently-labeled ATP analog termed deac-aminoATP to simulataneously visualize the stepping of fluorescently-labeled myosin V HMM and binding and dissociation of the nucleotide using the FIONA method. These studies show that there is a direct one to one coupling of myosin movement and nucleotide binding/dissociation and that there is strong gating between the kinetics of the two heads such that ADP release from the lead head is essentially prevented as long as the trail head is still attached.
| Andrecka, Joanna; Ortega Arroyo, Jaime; Takagi, Yasuharu et al. (2015) Structural dynamics of myosin 5 during processive motion revealed by interferometric scattering microscopy. Elife 4: |
| Ortega Arroyo, J; Andrecka, J; Spillane, K M et al. (2014) Label-free, all-optical detection, imaging, and tracking of a single protein. Nano Lett 14:2065-70 |
| Hammer 3rd, John A; Sellers, James R (2012) Walking to work: roles for class V myosins as cargo transporters. Nat Rev Mol Cell Biol 13:13-26 |
| Oke, Olusola A; Burgess, Stan A; Forgacs, Eva et al. (2010) Influence of lever structure on myosin 5a walking. Proc Natl Acad Sci U S A 107:2509-14 |
| Sellers, James R; Veigel, Claudia (2010) Direct observation of the myosin-Va power stroke and its reversal. Nat Struct Mol Biol 17:590-5 |
| Nagy, Nikolett T; Sakamoto, Takeshi; Takacs, Balazs et al. (2010) Functional adaptation of the switch-2 nucleotide sensor enables rapid processive translocation by myosin-5. FASEB J 24:4480-90 |
| Nagy, Attila; Piszczek, Grzegorz; Sellers, James R (2009) Extensibility of the extended tail domain of processive and nonprocessive myosin V molecules. Biophys J 97:3123-31 |
| Forgacs, Eva; Sakamoto, Takeshi; Cartwright, Suzanne et al. (2009) Switch 1 mutation S217A converts myosin V into a low duty ratio motor. J Biol Chem 284:2138-49 |
| Sellers, James R; Thirumurugan, Kavitha; Sakamoto, Takeshi et al. (2008) Calcium and cargoes as regulators of myosin 5a activity. Biochem Biophys Res Commun 369:176-81 |
| Takagi, Yasuharu; Yang, Yi; Fujiwara, Ikuko et al. (2008) Human myosin Vc is a low duty ratio, nonprocessive molecular motor. J Biol Chem 283:8527-37 |
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