The fastest‐actin‐based motor protein from the green algae, <i>Chara</i>, and its distinct mode of interaction with actin

Higashi-Fujime, Sugie; Ishikawa, Ryoki; Iwasawa, Hidehiro; Kagami, Osamu; Kurimoto, Eiji; Kohama, Kazuhiro; Hozumi, Tetsu

doi:10.1016/0014-5793(95)01208-v

Cited by 87 publications

(64 citation statements)

References 22 publications

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“…This motor can hydrolyze 400 molecules of ATP per second, which, combined with its long neck region , allows for translational speeds of 60 mm s 21 (Higashi-Fujime et al, 1995). These extremely fast turnover rates are consistent with the high speed of cytoplasmic streaming, around 60 mm s 21 , found in this organism (Kamiya and Kuroda, 1956), suggesting that myosin XI is responsible for this stunning example of intracellular motility.…”

Section: Not All Motors Are Created Equal: Differences In Enzymatic Psupporting

confidence: 65%

Update on Myosin Motors: Molecular Mechanisms and Physiological Functions

Ryan

Nebenführ

2017

Plant Physiol.

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Section: Not All Motors Are Created Equal: Differences In Enzymatic Psupporting

confidence: 65%

Update on Myosin Motors: Molecular Mechanisms and Physiological Functions

Ryan

Nebenführ

2017

Plant Physiol.

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“…Myosin purified from Chara corallina could translocate actin filaments in the in vitro motility assay at a velocity comparable to that of the cytoplasmic streaming (approximately 50 m s Ϫ1 ) (2,3). This velocity is about 10 times faster than that of the fast skeletal muscle myosin and the Chara myosin is the fastest motor protein known so far.…”

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confidence: 90%

Unique charge distribution in surface loops confers high velocity on the fast motor protein Chara myosin

Ito

Yamaguchi

Yanase

et al. 2009

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

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Most myosins have a positively charged loop 2 with a cluster of lysine residues that bind to the negatively charged N-terminal segment of actin. However, the net charge of loop 2 of very fast Chara myosin is zero and there is no lysine cluster in it. In contrast, Chara myosin has a highly positively charged loop 3. To elucidate the role of these unique surface loops of Chara myosin in its high velocity and high actin-activated ATPase activity, we have undertaken mutational analysis using recombinant Chara myosin motor domain. It was found that net positive charge in loop 3 affected Vmax and Kapp of actin activated ATPase activity, while it affected the velocity only slightly. The net positive charge in loop 2 affected Kapp and the velocity, although it did not affect Vmax. Our results suggested that Chara myosin has evolved to have highly positively charged loop 3 for its high ATPase activity and have less positively charged loop 2 for its high velocity. Since high positive charge in loop 3 and low positive charge in loop 2 seem to be one of the reasons for Chara myosin's high velocity, we manipulated charge contents in loops 2 and 3 of Dictyostelium myosin (class II). Removing positive charge from loop 2 and adding positive charge to loop 3 of Dictyostelium myosin made its velocity higher than that of the wild type, suggesting that the charge strategy in loops 2 and 3 is widely applicable.actin ͉ ATPase ͉ motility ͉ cytoplasmic streaming ͉ molecular engineering C ytoplasmic streaming in characean algal cells is extremely fast, and this streaming is brought about by the movement of myosin-coated organelles along actin filament bundles fixed inside the cell (1). Myosin purified from Chara corallina could translocate actin filaments in the in vitro motility assay at a velocity comparable to that of the cytoplasmic streaming (approximately 50 m s Ϫ1 ) (2, 3). This velocity is about 10 times faster than that of the fast skeletal muscle myosin and the Chara myosin is the fastest motor protein known so far. We have cloned cDNA of the Chara myosin heavy chain (4) and succeeded in expressing functional motor domain (5, 6). The velocity of the expressed motor domain measured by in vitro motility assay was comparable to that of the native Chara myosin if we consider the difference in the lever arm length. Relevant steps in actomyosin ATPase cycle to the sliding velocity are ADP release from actomyosin and ATP reassociation resulting in actin-myosin dissociation. Kinetic analyses of Chara myosin motor domain revealed that both the ADP release and the ATP-induced dissociation from actin were very fast (6). Time spent in the strongly bound state with actin, which was estimated from these rates, was less than 1 ms. This very short strongly bound state, together with a large step size (7), are probably the reason for the very high velocity of Chara myosin. Actin-activated ATPase activity of expressed Chara motor domain was approximately 500 s Ϫ1 head Ϫ1

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“…In the presence of Mg-ATP, muscle myosin-coated beads slide on actin cables with velocities similar to the maximum unloaded velocity of actin-myosin sliding in muscle (Sheetz and Spudich, 1983;Oiwa et al, 1990), whereas muscle actin filaments move on cytoplasmic myosin-coated glass surface with velocities similar to those of native cytoplasmic streaming (Higashi-Fujime et al, 1995). These results indicate that it is cytoplasmic myosin that is responsible for the rapid cytoplasmic streaming.…”

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confidence: 79%

“…Using the centrifuge microscope, Chaen et al (1995) showed that the Ca 2+ -induced actin-cytoplasmic myosin linkages are stronger than their rigor linkages, suggesting that a cytoplasmic myosin head can bind with actin at two different sites. Higashi-Fujime et al (1995) reported that proteolytic cleavage of actin impaired ATPdependent actin filament sliding on skeletal muscle myosin but not on cytoplasmic myosin, suggesting that cytoplasmic myosin may interact with actin at sites different from those of muscle myosin. …”

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confidence: 99%

Force–velocity relationships in actin–myosin interactions causing cytoplasmic streaming in algal cells

Sugi

Chaen

2003

Journal of Experimental Biology

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Cytoplasmic streaming is widely observed in a variety of plant cells. The mechanism of cytoplasmic streaming has been studied most intensively with giant internodal cells of green algae because of their large size (diameter, approximately 500·µm; length, >10·cm). It is now well established that cytoplasmic streaming is caused by ATP-dependent interaction between well-organized actin filament arrays fixed on the rows of chloroplasts (actin cables) and 'cytoplasmic' myosin molecules attached to amorphous cytoplasmic organelles. The sliding between actin and cytoplasmic myosin results in movement of the organelles in one direction, determined by the polarity of the actin cables, thus causing streaming of the surrounding cytoplasm (Kamiya and Kuroda, 1956;Kamitsubo, 1966; Nagai and Rebhum, 1966;Kersey and Wessels, 1976;Kato and Tonomura, 1977;Nagai and Hayama, 1979; Fig.·1). The direction of cytoplasmic streaming is reversed across the indifferent zone where chloroplasts are absent, reflecting the reversal of actin cable polarity across the indifferent zone. As the two indifferent zones run parallel to the cell's long axis, cytoplasmic streaming rotates longitudinally in the cell.Although both cytoplasmic streaming and muscle contraction are caused by ATP-dependent actin-myosin interaction, they differ from each other in some properties. Contraction and relaxation in skeletal muscle is regulated by changes in intracellular Ca 2+ concentration via regulatory proteins on actin filaments (Ebashi and Endo, 1968); a muscle is relaxed at pCa ≥7 and fully contracts at pCa ≤5. By contrast, cytoplasmic streaming occurs continuously even at pCa ≥7 and stops at pCa ≤6 (Williamson, 1975;Williamson and Ashley, 1982;Tominaga et al., 1983). Latex beads coated with skeletal muscle myosin slide along actin cables when they are introduced into an internodal cell with Mg-ATP, but the bead movement is insensitive to pCa (Shimmen and Yano, 1984), indicating that regulatory proteins are absent on actin cables, while cytoplasmic myosin is responsible for the Ca 2+ -induced stoppage of cytoplasmic streaming. Using the centrifuge microscope, Chaen et al. (1995) showed that the Ca 2+ -induced actin-cytoplasmic myosin linkages are stronger than their rigor linkages, suggesting that a cytoplasmic myosin head can bind with actin at two different sites. Higashi-Fujime et al. (1995) reported that proteolytic cleavage of actin impaired ATPdependent actin filament sliding on skeletal muscle myosin but not on cytoplasmic myosin, suggesting that cytoplasmic myosin may interact with actin at sites different from those of muscle myosin. 1971The Journal of Experimental Biology 206, 1971Biology 206, -1976Biology 206, © 2003 The Company of Biologists Ltd doi:10.1242/jeb.00239Cytoplasmic streaming in giant internodal cells of green algae is caused by ATP-dependent sliding between actin cables fixed on chloroplast rows and cytoplasmic myosin molecules attached to cytoplasmic organelles. Its velocity (≥50·µm·s -1 ) is many times larger than the maximum ...

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The fastest‐actin‐based motor protein from the green algae, Chara, and its distinct mode of interaction with actin

Cited by 87 publications

References 22 publications

Update on Myosin Motors: Molecular Mechanisms and Physiological Functions

Update on Myosin Motors: Molecular Mechanisms and Physiological Functions

Unique charge distribution in surface loops confers high velocity on the fast motor protein Chara myosin

Force–velocity relationships in actin–myosin interactions causing cytoplasmic streaming in algal cells

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