Influence of NaCl on the kinetic behaviour of mammalian muscle acetylcholinesterase

Area of Science:

• Biochemistry
• Enzymology
• Muscle Physiology

Background:

• Acetylcholinesterase (AChE) is crucial for neurotransmission.
• Understanding AChE's solubilization and kinetic properties in muscle is important for neuromuscular function studies.

Purpose of the Study:

• To investigate the solubilization of acetylcholinesterase from rabbit white muscle.
• To characterize the kinetic behavior of solubilized AChE, including cooperativity and the effect of salt concentration.

Main Methods:

• Solubilization of AChE using dilute buffer and Triton X-100.
• Enzyme activity assays at varying substrate and NaCl concentrations.
• Kinetic analysis to determine Km and cooperativity.

Main Results:

• AChE was effectively solubilized, with evidence of an occluded enzyme form in muscle.
• Triton X-100-solubilized AChE exhibited negative cooperativity at suboptimal substrate concentrations.
• Enzyme activity and Km were significantly affected by NaCl concentration, indicating salt's influence on substrate binding.

Conclusions:

• Rabbit white muscle contains a portion of acetylcholinesterase in an occluded state.
• The kinetic behavior of solubilized AChE is complex, involving negative cooperativity and sensitivity to ionic strength.
• A proposed kinetic model suggests substrate binding to both catalytic and peripheral anionic sites, with NaCl competing for these sites.