Light Chain Mutation Effects on the Dynamics of Thrombin.

Thrombin plays an important role in the process of hemostasis and blood coagulation. Studies in thrombin can help us find ways to treat cancer because thrombin is able to reduce the characteristic hypercoagulability of cancer. Thrombin is composed of two chains, the light chain and the heavy chain. The function of the heavy chain has been largely explored, while the function of the light chain was obscured until several disease-associated mutations in the light chain come to light. In this study, we want to explore the dynamic and conformation effects of mutations on the light chain further to determine possible associations between mutation, conformational changes, and disease. The study, which is a follow-up for our studies on apo thrombin and the mutant, ΔK9, mainly focuses on the mutants E8K and R4A. E8K is a disease-associated mutation, and R4A is used to study the role of Arg4, which is suggested experimentally to play a critical role for thrombin's catalytic activities. We performed five all-atom one microsecond-scale molecular dynamics (MD) simulations for both E8K and R4A, and quantified the changes in the conformational ensemble of the mutants. From the root-mean-square fluctuations (RMSF) for the α-carbons, we find that the atomic fluctuations change in the mutants in the 60s loop and γ loop. The correlation coefficients for the α-carbons indicate that the correlation relation for atom-pairs in the protein is also impacted. The clustering analysis and the principal component analysis (PCA) consistently tell us that the catalytic pocket and the regulatory loops are destabilized by the mutations. We also find that there are two binding modes for Na+ by clustering the vector difference between the Na+ ions and the 220s loop. After further analysis, we find that there is a relation between the Na+ binding and the rigidification of the γ loop, which may shed light on the mysterious role of the γ loop in thrombin.