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The reversibility of adenosine triphosphate cleavage by myosin
Abstract:
For the simplest kinetic model the reverse rate constants (k(-1) and k(-2)) associated with ATP binding and cleavage on purified heavy meromyosin and heavy meromyosin subfragment 1 from rabbit skeletal muscle in the presence of 5mm-MgCl(2), 50mm-KCl and 20mm-Tris-HCl buffer at pH8.0 and 22 degrees C are: k(-1)<0.02s(-1) and k(-1)=16s(-1). Apparently, higher values of k(-1) and k(-2) are found with less-purified protein preparations. The values of k(-1) and k(-2) satisfy conditions required by previous (18)O-incorporation studies of H(2) (18)O into the P(i) moiety on ATP hydrolysis and suggest that the cleavage step does involve hydrolysis of ATP or formation of an adduct between ATP and water. The equilibrium constant for the cleavage step at the myosin active site is 9. If the cycle of events during muscle contraction is described by the model proposed by Lymn & Taylor (1971), the fact that there is only a small negative standard free-energy change for the cleavage step is advantageous for efficient chemical to mechanical energy exchange during muscle contraction.
Insights
This study determined reverse rate constants for ATP binding and cleavage in rabbit skeletal muscle myosin. These findings support ATP hydrolysis during muscle contraction and efficient energy conversion.
Area of Science:
- Biochemistry
- Muscle Physiology
- Enzymology
Background:
- Muscle contraction relies on the biochemical cycle of myosin interaction with actin.
- Understanding the kinetics of ATP hydrolysis by myosin is crucial for elucidating energy transduction mechanisms.
Purpose of the Study:
- To determine the reverse rate constants (k(-1) and k(-2)) for ATP binding and cleavage by heavy meromyosin and heavy meromyosin subfragment 1.
- To investigate the implications of these rate constants for the mechanism of ATP hydrolysis and energy conversion during muscle contraction.
Main Methods:
- Kinetic analysis of purified heavy meromyosin and heavy meromyosin subfragment 1 from rabbit skeletal muscle.
- Measurement of reverse rate constants under specific buffer conditions (5mM MgCl2, 50mM KCl, 20mM Tris-HCl, pH 8.0, 22°C).
Main Results:
- Reverse rate constants were determined: k(-1)<0.02s(-1) and k(-1)=16s(-1).
- Higher rate constants were observed with less purified protein preparations.
- The equilibrium constant for the cleavage step at the myosin active site was found to be 9.
- Results are consistent with previous (18)O-incorporation studies, suggesting ATP hydrolysis or adduct formation.
Conclusions:
- The determined rate constants support a mechanism involving ATP hydrolysis or adduct formation during the cleavage step.
- A small negative standard free-energy change for cleavage is advantageous for efficient chemical to mechanical energy exchange in muscle contraction, as per the Lymn & Taylor model.