Observing Dynamic Conformational Changes within the Coiled-Coil Domain of Different Laminin Isoforms Using High-Speed...

Area of Science:

• Biochemistry
• Structural Biology
• Cell Biology

Background:

• Laminins are essential extracellular matrix glycoproteins involved in cell adhesion and tissue organization.
• Their cross-shaped structure comprises α, ß, and γ-chains, with laminin G-like (LG) domains at the C-terminus mediating cell binding.
• Previous studies suggested dynamic conformational changes regulate laminin function, but lacked single-molecule dynamic analysis.

Purpose of the Study:

• To investigate the dynamic conformational changes of laminin-111 and laminin-332 at the single-molecule level.
• To characterize the ultrastructure and conformational flexibility of laminin coiled-coil and LG domains under physiological conditions.
• To provide novel insights into laminin molecular dynamics using high-speed atomic force microscopy.

Main Methods:

• High-speed atomic force microscopy (HS-AFM) was employed for real-time, sub-second imaging of laminin-111 and laminin-332.
• Surface immobilization and imaging conditions were optimized for high-resolution visualization.
• Complementary computational methods including AlphaFold structure prediction and molecular dynamics simulations were utilized.

Main Results:

• Laminin-111 exhibited a stable, S-shaped coiled-coil domain with minimal conformational changes.
• Laminin-332 displayed rapid switching of its coiled-coil domain between straight and bent conformations around a central hinge.
• Dynamic rearrangements of C-terminal LG domains between compact and open states were observed for both laminin isoforms.

Conclusions:

• HS-AFM directly visualizes dynamic molecular rearrangements in laminin isoforms, offering unprecedented structural insights.
• Laminin-332 possesses a flexible coiled-coil domain, suggesting a distinct mechanism for regulating its function compared to laminin-111.
• The study highlights the importance of dynamic conformational changes in laminin function and provides a powerful technique for their analysis.