Abstract
Blood coagulation factor V (FV) is activated either by Factor X or thrombin, cleaving at three different sites viz., Site I (Arg709-Ser710), site II (Arg1018-Thr1019), and site III (Arg1545-Ser1546). Russell's viper venom factor V activator (RVV-V) is a thrombin-like serine proteinase that activates FV with selective, single cleavage at site III. A long lasting effort is being pending in understanding the 'selective' binding specificity of the RVV-V towards site III. Here, we present the binding kinetic study of RVV-V with two designed peptides corresponding to the regions from site I (Gln699-Asn713) and site II (1008Lys-Pro1022), respectively, that include 15 amino acids. Our investigation for justifying the binding efficacy and kinetics of peptides includes SPR method, protein-peptide docking, molecular dynamics simulation, and principal component analysis (PCA). Surprisingly, the SPR experiment disclosed that the Peptide II showed a lower binding affinity with KD of 2.775 mM while the Peptide I showed none. Docking and simulation of both the peptides with RVV-V engaged either rooted or shallow binding for Peptide II and Peptide I respectively. The peptide binding resulted in global conformational changes in the native fold of RVV-V, whereas the similar studies for thrombin failed to make major changes in the native fold. In support, the PCA analysis for RVV-V showed the dislocation of catalytic triad upon binding both the peptides. Hence, RVV-V, a serine protease, is incompetent in cleaving these two sites. This study suggests a transition in RVV-V from the native rigid to the distorted flexible structure and paves a way to design a new peptide substrate/inhibitor.