Simulations suggest double sodium binding induces unexpected conformational changes in thrombin

Thrombin is a multifunctional Na+-activated protein. There is one Na+-binding site that has been the focus of the study of the thrombin. Previous work based on molecular dynamics (MD) simulations suggests that there are two Na+-binding sites in thrombin. This work also suggests that thrombin exhibits dynamic, aka generalized, allostery. These sites can be bound separately or simultaneously. In this study, we performed 12 independent 2μs all-atom MD simulations of wild-type (WT) thrombin and studied the different Na+binding modes. We used root-mean-square fluctuations to study how different binding events rigidify different regions. We also used correlation matrixes to suggest regions that may play a role in thrombin's allosteric response. Non-parametric clustering and principal component analyses are used to examine the conformations induced by different sodium binding modes. Based on these analyses, as well as SASA analysis, we suggest that the double binding mode might be an inactive mode. We also suggest that the kinetic scheme for Na+ binding to thrombin involves a multiple-step mechanism.