Real-time visualization of conformational changes within single MloK1 cyclic nucleotide-modulated channels

Area of Science:

• Ion channel biophysics
• Molecular biology
• Structural biology

Background:

• Cyclic nucleotide-modulated (CNM) ion channels are crucial for cellular signaling.
• Understanding the dynamic conformational changes during channel gating is essential for elucidating their function.
• Previous studies lacked real-time imaging of these dynamic processes.

Purpose of the Study:

• To visualize real-time, ligand-induced conformational changes in single CNM channels.
• To investigate the dynamic behavior of the Mesorhizobium loti K+ channel (MloK1) during gating.
• To correlate conformational states with channel activity.

Main Methods:

• High-speed atomic force microscopy (HS-AFM) was employed for real-time imaging.
• Single MloK1 channels were studied in the presence and absence of cyclic adenosine monophosphate (cAMP).
• Analysis focused on conformational transitions and blinking dynamics.

Main Results:

• In the presence of cAMP, MloK1 channels predominantly adopted a stable conformation.
• A subset of channels exhibited dynamic blinking between distinct conformations with varying heights.
• cAMP depletion led to increased blinking and altered blinking heights, indicating ligand dissociation.

Conclusions:

• MloK1 transitions between mobile conformations (ligand-absent) and a stable conformation (ligand-bound) during gating.
• These conformational dynamics are central to the pore gating mechanism of CNM channels.
• HS-AFM provides unprecedented insight into the real-time molecular mechanisms of ion channel function.