Thrombosis and haemostasis vol:100 issue:6 pages:1058-1067
Multimerin I (MMRNI) is a polymeric, factorV (FV) binding protein that is stored in platelet and endothelial cell secretion granules but is undetectable in normal plasma. In human platelet alpha-granules, FV is stored complexed to MMRNI, predominantly by noncovalent binding interactions. The FV binding site for MMRNI is located in the light chain, where it overlaps the C1 and C2 domain membrane binding sites essential for activated FV (FVa) procoagulant function. Surface plasmon resonance (SPR), circular dichroism (CID) and thrombin generation assays were used to study the binding of FV and FVa to MMRNI, and the functional consequences. FV and FVa bound MMRNI with high affinities (K-D:2 and 7 nM, respectively). FV dissociated more slowly from MMRNI than FVa in SPR experiments, and CD analyses suggested greater conformational changes in mixtures of FV and MMRNI than in mixtures of FVa and MMRNI. SPR analyses indicated that soluble phosphatidylserine (1,2-Dicaproylsn-glycero-3-phospho-L-serine) competitively inhibited both FV-MMRNI and FVa-MMRNI binding. Furthermore, exogenous MMRNI delayed and reduced thrombin generation by plasma and platelets, and it reduced thrombin generation by preformed FVa. Exogenous MMRNI also delayed FV activation, triggered by adding tissue factor to plasma, or by adding purified thrombin or factor Xa to purified FV. The high affinity binding of FV to MMRNI may facilitate the costorage of the two proteins in platelet alpha-granules. As a consequence, MMRNI release during platelet activation may limit platelet dependent thrombin generation in vivo.