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Biophys J. 2008 Dec 15;95(12):5798-808. doi: 10.1529/biophysj.108.130435. Epub 2008 Oct 3.

Effect of inorganic phosphate on the force and number of myosin cross-bridges during the isometric contraction of permeabilized muscle fibers from rabbit psoas.

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Laboratorio di Fisiologia, DBE, and CNISM, Università di Firenze, Florence, Italy.


The relation between the chemical and mechanical steps of the myosin-actin ATPase reaction that leads to generation of isometric force in fast skeletal muscle was investigated in demembranated fibers of rabbit psoas muscle by determining the effect of the concentration of inorganic phosphate (Pi) on the stiffness of the half-sarcomere (hs) during transient and steady-state conditions of the isometric contraction (temperature 12 degrees C, sarcomere length 2.5 mum). Changes in the hs strain were measured by imposing length steps or small 4 kHz oscillations on the fibers in control solution (without added Pi) and in solution with 3-20 mM added Pi. At the plateau of the isometric contraction in control solution, the hs stiffness is 22.8 +/- 1.1 kPa nm(-1). Taking the filament compliance into account, the total stiffness of the array of myosin cross-bridges in the hs (e) is 40.7 +/- 3.7 kPa nm(-1). An increase in [Pi] decreases the stiffness of the cross-bridge array in proportion to the isometric force, indicating that the force of the cross-bridge remains constant independently of [Pi]. The rate constant of isometric force development after a period of unloaded shortening (r(F)) is 23.5 +/- 1.0 s(-1) in control solution and increases monotonically with [Pi], attaining a maximum value of 48.6 +/- 0.9 s(-1) at 20 mM [Pi], in agreement with the idea that Pi release is a relatively fast step after force generation by the myosin cross-bridge. During isometric force development at any [Pi], e and thus the number of attached cross-bridges increase in proportion to the force, indicating that, independently of the speed of the process that leads to myosin attachment to actin, there is no significant (>1 ms) delay between generation of stiffness and generation of force by the cross-bridges.

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