In silico conformational dynamics of the α-actinin-2 actin-binding domain upon phosphorylation

Area of Science:

• Cardiovascular Biology
• Molecular and Structural Biology
• Biophysics

Background:

• α-actinin-2 phosphorylation in cardiomyocytes increases with mechanical stress, aiding force adaptation.
• The structural impact of these phosphorylation sites on F-actin binding and sarcomere assembly remains unclear.

Purpose of the Study:

• To elucidate the mechanisms by which α-actinin-2 regulates sarcomere assembly dynamics in the heart.
• To investigate phosphorylation-specific structural modifications within the α-actinin-2 actin-binding domain (ABD).

Main Methods:

• Structural modeling of phosphorylation sites (T43, S50, S147, T237) and phosphomimetic variants (T43D, S50D, S147D, T237D) using AlphaFold 3.
• Molecular dynamics (MD) simulations to quantify conformational changes and electrostatic interactions at the ABD.
• Analysis of changes in domain distances, torsion angles, and electrostatic potentials.

Main Results:

• Phosphorylation and phosphomimetic mutations in α-actinin-2 ABD increased the distance between calponin homology domains (CH1 and CH2), favoring an 'open' ABD conformation.
• These structural changes correlated with improved electrostatic interactions between α-actinin-2 and F-actin.
• MD simulations indicated that phosphorylation enhances conformational flexibility, particularly at sites S147D, T237D, and T43D.

Conclusions:

• α-actinin-2 phosphorylation destabilizes the closed ABD conformation, promoting an open state conducive to F-actin binding.
• This phosphorylation-driven mechanism is critical for regulating sarcomere assembly and stabilization in response to mechanical forces in the heart.