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Electron microscopic recording of myosin head power stroke in hydrated myosin filaments.

Sugi H, Chaen S, Akimoto T, Minoda H, Miyakawa T, Miyauchi Y, Tanokura M, Sugiura S - Sci Rep (2015)

Bottom Line: Using the gas environmental chamber, we have succeeded in recording the power stroke of position-marked myosin heads in hydrated mixture of actin and myosin filaments in a nearly isometric condition, in which myosin heads do not produce gross myofilament sliding, but only stretch adjacent elastic structures.After exhaustion of applied ATP, individual myosin heads return towards their initial position.The advantages of the present study over other in vitro motility assay systems, using myosin heads detached from myosin filaments, are discussed.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, School of Medicine, Teikyo University, Itabashi-ku, Tokyo 173-8605, Japan.

ABSTRACT
Muscle contraction results from cyclic attachment and detachment between myosin heads and actin filaments, coupled with ATP hydrolysis. Despite extensive studies, however, the amplitude of myosin head power stroke still remains to be a mystery. Using the gas environmental chamber, we have succeeded in recording the power stroke of position-marked myosin heads in hydrated mixture of actin and myosin filaments in a nearly isometric condition, in which myosin heads do not produce gross myofilament sliding, but only stretch adjacent elastic structures. On application of ATP, individual myosin heads move by ~3.3 nm at the distal region, and by ~2.5 nm at the proximal region of myosin head catalytic domain. After exhaustion of applied ATP, individual myosin heads return towards their initial position. At low ionic strength, the amplitude of myosin head power stroke increases to >4 nm at both distal and proximal regions of myosin heads catalytic domain, being consistent with the report that the force generated by individual myosin heads in muscle fibers is enhanced at low ionic strength. The advantages of the present study over other in vitro motility assay systems, using myosin heads detached from myosin filaments, are discussed.

No MeSH data available.


Related in: MedlinePlus

Histograms showing the amplitude distribution of ATP-induced myosin head power stroke at external KCl concentration of 20 mM.Individual myosin heads were position-marked with antibody 1 at the distal region of myosin head CAD in (A), and with antibody 2 at the proximal region of myosin head CAD in (B).
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f5: Histograms showing the amplitude distribution of ATP-induced myosin head power stroke at external KCl concentration of 20 mM.Individual myosin heads were position-marked with antibody 1 at the distal region of myosin head CAD in (A), and with antibody 2 at the proximal region of myosin head CAD in (B).

Mentions: To test the validity of this expectation, we performed experiments, in which the KCl concentration of experimental solution surrounding the filament mixture was lowered from 120 to 20 mM (corresponding to a reduction of ionic strength μ from 170 to 50 mM). Examples of IP records taken before and after ATP application at low ionic strength are shown in Fig. 4. In this particular case, the IP record of gold particles attached to individual myosin heads, taken before ATP application is colored red (A), and that taken after ATP application is colored blue (B), while the bottom diagram (C) shows ATP-induced change in position of each gold particle, i.e. the ATP-induced power stroke in individual myosin heads. The amplitude of ATP-induced myosin head power stroke was found to increase markedly by the reduction of ionic strength. In contrast with the histograms shown in Fig. 3, the histograms of myosin head power stroke at low ionic strength exhibited distinct peaks at 2.5–5 nm around both distal and proximal regions of myosin head catalytic domain (Fig. 5A,B). The average amplitude of myosin head power stroke at low ionic strength was 4.4 ± 0.1 nm (mean ± SD, n = 361) and 4.3 ± 0.2 nm (mean ± SD, n = 305) at the distal and at the proximal regions of myosin head catalytic domain, respectively, indicating that the amplitude of myosin head power stroke does not differ significantly between the distal and the proximal regions of myosin head catalytic domain at low ionic strength.


Electron microscopic recording of myosin head power stroke in hydrated myosin filaments.

Sugi H, Chaen S, Akimoto T, Minoda H, Miyakawa T, Miyauchi Y, Tanokura M, Sugiura S - Sci Rep (2015)

Histograms showing the amplitude distribution of ATP-induced myosin head power stroke at external KCl concentration of 20 mM.Individual myosin heads were position-marked with antibody 1 at the distal region of myosin head CAD in (A), and with antibody 2 at the proximal region of myosin head CAD in (B).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4620508&req=5

f5: Histograms showing the amplitude distribution of ATP-induced myosin head power stroke at external KCl concentration of 20 mM.Individual myosin heads were position-marked with antibody 1 at the distal region of myosin head CAD in (A), and with antibody 2 at the proximal region of myosin head CAD in (B).
Mentions: To test the validity of this expectation, we performed experiments, in which the KCl concentration of experimental solution surrounding the filament mixture was lowered from 120 to 20 mM (corresponding to a reduction of ionic strength μ from 170 to 50 mM). Examples of IP records taken before and after ATP application at low ionic strength are shown in Fig. 4. In this particular case, the IP record of gold particles attached to individual myosin heads, taken before ATP application is colored red (A), and that taken after ATP application is colored blue (B), while the bottom diagram (C) shows ATP-induced change in position of each gold particle, i.e. the ATP-induced power stroke in individual myosin heads. The amplitude of ATP-induced myosin head power stroke was found to increase markedly by the reduction of ionic strength. In contrast with the histograms shown in Fig. 3, the histograms of myosin head power stroke at low ionic strength exhibited distinct peaks at 2.5–5 nm around both distal and proximal regions of myosin head catalytic domain (Fig. 5A,B). The average amplitude of myosin head power stroke at low ionic strength was 4.4 ± 0.1 nm (mean ± SD, n = 361) and 4.3 ± 0.2 nm (mean ± SD, n = 305) at the distal and at the proximal regions of myosin head catalytic domain, respectively, indicating that the amplitude of myosin head power stroke does not differ significantly between the distal and the proximal regions of myosin head catalytic domain at low ionic strength.

Bottom Line: Using the gas environmental chamber, we have succeeded in recording the power stroke of position-marked myosin heads in hydrated mixture of actin and myosin filaments in a nearly isometric condition, in which myosin heads do not produce gross myofilament sliding, but only stretch adjacent elastic structures.After exhaustion of applied ATP, individual myosin heads return towards their initial position.The advantages of the present study over other in vitro motility assay systems, using myosin heads detached from myosin filaments, are discussed.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, School of Medicine, Teikyo University, Itabashi-ku, Tokyo 173-8605, Japan.

ABSTRACT
Muscle contraction results from cyclic attachment and detachment between myosin heads and actin filaments, coupled with ATP hydrolysis. Despite extensive studies, however, the amplitude of myosin head power stroke still remains to be a mystery. Using the gas environmental chamber, we have succeeded in recording the power stroke of position-marked myosin heads in hydrated mixture of actin and myosin filaments in a nearly isometric condition, in which myosin heads do not produce gross myofilament sliding, but only stretch adjacent elastic structures. On application of ATP, individual myosin heads move by ~3.3 nm at the distal region, and by ~2.5 nm at the proximal region of myosin head catalytic domain. After exhaustion of applied ATP, individual myosin heads return towards their initial position. At low ionic strength, the amplitude of myosin head power stroke increases to >4 nm at both distal and proximal regions of myosin heads catalytic domain, being consistent with the report that the force generated by individual myosin heads in muscle fibers is enhanced at low ionic strength. The advantages of the present study over other in vitro motility assay systems, using myosin heads detached from myosin filaments, are discussed.

No MeSH data available.


Related in: MedlinePlus