Light Chain Mutation Effects on the Dynamics of Thrombin

Area of Science:

• Biochemistry and Molecular Biology
• Protein Dynamics and Mutagenesis

Background:

• Thrombin is crucial for hemostasis and cancer hypercoagulability.
• The thrombin light chain's function is less understood than its heavy chain.
• Disease-associated mutations in the light chain highlight its importance.

Purpose of the Study:

• To investigate the dynamic and conformational effects of light chain mutations in thrombin.
• To explore the link between mutation, conformational changes, and disease.
• To analyze the impact of E8K (disease-associated) and R4A (catalytic role) mutations.

Main Methods:

• Performed five all-atom, microsecond-scale molecular dynamics (MD) simulations for E8K and R4A mutants.
• Quantified conformational ensemble changes using root-mean-square fluctuations (RMSF) and correlation coefficients.
• Utilized clustering and principal component analysis (PCA) to assess protein structure and dynamics.

Main Results:

• Mutations E8K and R4A altered atomic fluctuations in the 60s and γ loops.
• Protein atom-pair correlations were impacted, and PCA/clustering revealed destabilization of the catalytic pocket and regulatory loops.
• Identified two Na+ binding modes, linking Na+ binding to the rigidification of the γ loop.

Conclusions:

• Thrombin light chain mutations significantly alter protein dynamics and conformation.
• These changes destabilize key functional regions, including the catalytic pocket.
• The study provides insights into the role of the γ loop and Na+ binding in thrombin function.