Mechanism of Cardiac Troponin C Calcium Sensitivity Modulation by Small Molecules Illuminated by Umbrella Sampling...

Area of Science:

• Biochemistry
• Molecular Biology
• Cardiovascular Research

Background:

• Cardiac troponin C (cTnC) is crucial for muscle contraction by binding calcium and cardiac troponin I (cTnI).
• Dysfunctional muscle contraction, often due to heritable cardiomyopathies, makes cTnC a therapeutic target.
• Existing structural studies identify small molecule binding sites but not their energetic influence on cTnC's regulatory domain opening.

Purpose of the Study:

• To investigate the impact of calcium sensitivity modulators on the free energy of cTnC regulatory domain opening.
• To elucidate the mechanisms by which small molecules influence cTnC function.
• To apply umbrella sampling to study molecular interactions with cTnC.

Main Methods:

• Umbrella sampling simulations were employed to calculate the free energy landscape of cTnC regulatory domain opening.
• Conventional molecular dynamics (MD) simulations were used to assess the binding stability of small molecules.
• The effects of bepridil, dfbp-o, 3-methyldiphenylamine (DPA), and W7 on cTnC energetics were analyzed.

Main Results:

• All tested small molecules reduced the free energy of cTnC opening in the absence of cTnI.
• DPA and dfbp-o stabilized the open cTnC conformation in the presence of cTnI.
• Bepridil and W7 destabilized the open conformation when cTnI was present, while NSC34337 showed poor binding.

Conclusions:

• This study introduces a novel umbrella sampling approach to quantify small molecule effects on cTnC.
• Small molecules differentially modulate cTnC's open conformation energetics, influencing calcium sensitivity.
• Effective calcium sensitivity modulators likely stabilize the open hydrophobic patch conformation of cTnC.