The light chain composition of chicken brain myosin‐Va: Calmodulin, myosin‐II essential light chains, and 8‐kDa dynein light chain/PIN

Espíndola, Foued Salmen; Suter, Daniel M.; Partata, Leticia B.E.; Cao, Tracy T.; Wolenski, Joseph S.; Cheney, Richard E.; King, Stephen M.; Mooseker, Mark S.

doi:10.1002/1097-0169(200012)47:4<269::aid-cm2>3.3.co;2-7

Cited by 47 publications

(53 citation statements)

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“…We also coexpressed non-muscle myosin essential light chain (LC17b) along with myosin Vb heavy chain and calmodulin, but we could not detect the essential light chain associated with the purified myosin Vb (data not shown). While chicken myosin Va purified from brain has two essential light chains (LC1sa and LC17b) in addition to calmodulin (40), mouse myosin Va purified from brain does not show the presence of any essential light chains (41). The result suggests that mammalian myosin Vb has calmodulin but not the essential light chains such as mammalian myosin Va. Figure 2B shows electron micrographs of M5BS1 and M5BHMM.…”

Section: Expression and Purification Of M5b Constructsmentioning

confidence: 98%

Mechanoenzymatic Characterization of Human Myosin Vb

et al. 2006

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There are three isoforms of class V myosin in mammals. While myosin Va has been studied well, little is known about the function of other myosin V isoforms (Vb and Vc) at a molecular level. Here we report the mechanoenzymatic function of human myosin Vb (HuM5B) for the first time. Electron microscopic observation showed that HuM5B has a double-headed structure with a long neck like myosin Va. V max and K actin of the actin-activated ATPase activity of HuM5B were 9.7 ( 0.4 s -1 and 8.5 ( 0.1 µM, respectively. K actin and K ATP of the actin-activated ATPase activity were significantly higher than those of myosin Va. ADP markedly inhibited the ATPase activity. The rate of release of ADP from acto-HuM5B was 12.2 ( 0.5 s -1 , which was comparable to the V max of the actin-activated ATPase activity. These results suggest that ADP release is the rate-limiting step for the actin-activated ATPase cycle; thus, HuM5B is a high duty ratio myosin. Consistently, the actin gliding velocity (0.22 ( 0.03 µm/s) remained constant at a low motor density. The actin filament landing assay revealed that a single HuM5B molecule is sufficient to move the actin filament continuously, indicating that HuM5b is a processive motor.Myosin is a mechanoenzymatic protein that interacts with actin and converts the chemical energy from ATP hydrolysis to the mechanical work. In addition to well-characterized conventional, filament-forming, two-headed myosin II of muscle and non-muscle cells, a number of myosin-like proteins have been discovered, and it is known that myosin constitutes a diverse superfamily divided into at least 18 classes (1-7). Class V myosins are one of the most ancient myosins of the superfamily, and their genes have been identified in species from yeast, Caenorhabditis elegans, and Drosophila to humans (7). Dictyostelium MyoJ and plant class XI and VIII myosins are structurally and functionally related to class V myosins. Yeast contains two class V myosins, while C. elegans and Drosophila contain a single class V myosin. In mammals, there are three distinct subclasses of myosin V. Mammalian class V myosin was first discovered from dilute mice that have a lighter color coat due to the lack of proper pigment transport (8). This myosin V is now classified as myosin Va, and most of the biochemical and cell biological works for class V myosin have been done with this myosin V isoform (see ref 9 for a review). There are two additional myosin V isoforms reported, i.e., myosin Vb and myosin Vc (10, 11), yet their properties at a molecular level have not been studied.The structure of myosin Va was visualized by electron microscopy, and it was shown that myosin Va is the twoheaded structure with the long neck connecting the head and the coiled-coil domain. On the basis of the amino acid sequence homology with myosin Va (10, 11), it is thought that, overall, the structure of myosin Vb and myosin Vc resembles that of myosin Va. The heavy chain of myosin Vb has a molecular mass of 214 kDa and composed of the N-terminal conserved m...

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Section: Expression and Purification Of M5b Constructsmentioning

confidence: 98%

Mechanoenzymatic Characterization of Human Myosin Vb

et al. 2006

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“…17,18 This domain likely acts as a lever arm that amplifies small nucleotidedependent conformational changes in the motor domain to a large displacement. 19,20 Myosin V walks toward the plus end of actin by taking ∼37 nm steps per ATP hydrolyzed.…”

Section: Molecular Motorsmentioning

confidence: 99%

Fluorescence Imaging with One Nanometer Accuracy: Application to Molecular Motors

2005

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We introduce the technique of FIONA, fluorescence imaging with one nanometer accuracy. This is a fluorescence technique that is able to localize the position of a single dye within ∼1 nm in the x-y plane. It is done simply by taking the point spread function of a single fluorophore excited with wide field illumination and locating the center of the fluorescent spot by a two-dimensional Gaussian fit. We motivate the development of FIONA by unraveling the walking mechanism of the molecular motors myosin V, myosin VI, and kinesin. We find that they all walk in a hand-over-hand fashion.

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“…The function of DLC is regulated by PAK phosphorylation on DLC-Ser88 [157] that enhances endosomal trafficking and macropinocytosis [158]. Apart from its well known role as a component of dynein motors, DLC is also present as a stoichiometric subunit of the myosin V motors that transport short-range vesicles in the cell cortex [159,160] raising the possibility that PAK regulation of DLC function may affect traffic through both filament systems. Furthermore, like MP1/p14, LC8 (DLC1) also interacts with PAK1, a kinase that phosphorylates both MEK1 and LC8 [157].…”

Section: Erk and Microtubule And Motor Dynamicsmentioning

confidence: 99%

Scaffold mediated regulation of MAPK signaling and cytoskeletal dynamics: A perspective

Pullikuth

Catling

2007

Cellular Signalling

136

107

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Cell migration is critical for many physiological processes and is often misregulated in developmental disorders and pathological conditions including cancer and neurodegeneration. MAPK signaling and the Rho family of proteins are known regulators of cell migration that exert their influence on cellular cytoskeleton during cell adhesion and migration. Here we review data supporting the view that localized ERK signaling mediated through recently identified scaffold proteins may regulate cell migration.

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The light chain composition of chicken brain myosin‐Va: Calmodulin, myosin‐II essential light chains, and 8‐kDa dynein light chain/PIN

Cited by 47 publications

References 0 publications

Mechanoenzymatic Characterization of Human Myosin Vb

Mechanoenzymatic Characterization of Human Myosin Vb

Fluorescence Imaging with One Nanometer Accuracy: Application to Molecular Motors

Scaffold mediated regulation of MAPK signaling and cytoskeletal dynamics: A perspective

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