Kinetics of adenosine triphosphate hydrolysis by shortening myofibrils from rabbit psoas muscle.

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Abstract
1. Using a solenoid‐operated mixing device, time‐resolved measurements were made of shortening and accompanying ATP hydrolysis at 20 degrees C by myofibrils prepared from rabbit psoas muscle. 2. The extent of ATP hydrolysis was determined by an improved Malachite Green method for determination of inorganic phosphate (Pi) in the presence of a large excess of ATP. For the measurement of the change in sarcomere length by phase contrast microscopy, shortening was terminated without delay and artifact by a mixture of 0.2 M‐acetate (pH 4.6) and 1.25% (v/v) glutaraldehyde. 3. The shortening velocity per half‐sarcomere was 10 microns s‐1 in 25 mM‐KCl for sarcomere lengths above 1.4 microns, and at least 12 microns s‐1 in 150 mM‐KCl for sarcomere lengths above 1.7 microns. During this rapid shortening, there was no significant ATP turnover by cross‐bridges (upper 95% confidence limit: 0.14 mol (mol of myosin head)‐1 in 25 mM‐KCl; 0.12 mol mol‐1 in KCl solutions greater than or equal to 100 mM). 4. When the sarcomeres shortened below 1.7 microns in KCl concentrations greater than 100 mM or below 1.4 microns in 25 mM‐KCl, there was a transient acceleration of ATP hydrolysis (delayed ATP hydrolysis), which was then followed by a steady slow hydrolysis. 5. The magnitudes (+/‐ estimated standard deviation) of delayed ATP hydrolysis by myofibrils of initial sarcomere length 2.4 microns were 0.42 +/‐ 0.19, 0.31 +/‐ 0.10 and 0.17 +/‐ 0.09 mol (mol myosin head)‐1 in 25 mM, 100 mM and 150 mM‐KCl, respectively. For myofibrils of sarcomere length 2.0 microns, however, it decreased to 0.24 +/‐ 0.10 mol mol‐1 in 25 mM‐KCl or to an insignificant level in 150 mM‐KCl. 6. These results indicate that most of the ATP hydrolysis products remain bound to cross‐bridges during rapid shortening, and that when the force opposing shortening increases, a proportion of cross‐bridges rapidly dissociate the products and enter the next ATP cycle, which diminishes with the decrease in shortening distance as well as the increase in ionic strength. Such behaviour of the cross‐bridge is probably a manifestation of its energetic and kinetic properties in the state with bound ADP and Pi interacting with actin filaments at zero load and at a transition from zero to non‐zero loads.