Filamentous smooth muscle myosin is regulated by phosphorylation

Area of Science:

• Biochemistry
• Molecular Biology
• Muscle Physiology

Background:

• Determining enzymatic activity of filamentous dephosphorylated smooth muscle myosin is challenging due to MgATP-induced disassembly.
• Smooth muscle myosin's assembly state influences its enzymatic properties.

Purpose of the Study:

• To investigate the role of light chain phosphorylation in regulating smooth muscle myosin ATPase activity.
• To overcome challenges in studying dephosphorylated filamentous myosin.

Main Methods:

• Utilized monoclonal antirod antibodies to stabilize dephosphorylated myosin in its filamentous state.
• Performed steady-state and single-turnover actin-activated ATPase assays.
• Compared enzymatic activity of phosphorylated and antibody-stabilized dephosphorylated myosin filaments.

Main Results:

• Phosphorylated smooth muscle myosin filaments exhibited 30-100-fold higher actin-activated ATPase activity than dephosphorylated filaments.
• Dephosphorylated filaments and heavy meromyosin showed similar low rates of phosphate release, with or without actin.
• Antibody stabilization allowed measurement of dephosphorylated filamentous myosin's low basal activity.

Conclusions:

• Smooth muscle myosin light chain phosphorylation is a key regulatory mechanism for ATPase activity, independent of assembly state changes.
• Phosphorylation acts as an effective switch to control smooth muscle myosin filament activity.
• The regulatory potential of phosphorylation may exceed 100-fold.