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The roles of factor Va and protein S in formation of the activated protein C/protein S/factor Va inactivation complex
Author(s) -
Gierula Magdalena,
SallesCrawley Isabelle I.,
Santamaria Salvatore,
TerazOrosz Adrienn,
Crawley James T. B.,
Lane David A.,
Ahnström Josefin
Publication year - 2019
Publication title -
journal of thrombosis and haemostasis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.947
H-Index - 178
eISSN - 1538-7836
pISSN - 1538-7933
DOI - 10.1111/jth.14594
Subject(s) - chemistry , protein c , protein s , phospholipid , plasma protein binding , biochemistry , microbiology and biotechnology , biology , membrane
Abstract Background Activated protein C (APC)‐mediated inactivation of factor (F)Va is greatly enhanced by protein S. For inactivation to occur, a trimolecular complex among FVa, APC, and protein S must form on the phospholipid membrane. However, direct demonstration of complex formation has proven elusive. Objectives To elucidate the nature of the phospholipid‐dependent interactions among APC, protein S, and FVa. Methods We evaluated binding of active site blocked APC to phospholipid‐coated magnetic beads in the presence and absence of protein S and/or FVa. The importance of protein S and FV residues were evaluated functionally. Results Activated protein C alone bound weakly to phospholipids. Protein S mildly enhanced APC binding to phospholipid surfaces, whereas FVa did not. However, FVa together with protein S enhanced APC binding (>14‐fold), demonstrating formation of an APC/protein S/FVa complex. C4b binding protein‐bound protein S failed to enhance APC binding, agreeing with its reduced APC cofactor function. Protein S variants (E36A and D95A) with reduced APC cofactor function exhibited essentially normal augmentation of APC binding to phospholipids, but diminished APC/protein S/FVa complex formation, suggesting involvement in interactions dependent upon FVa. Similarly, FVa Nara (W1920R), an APC‐resistant FV variant, also did not efficiently incorporate into the trimolecular complex as efficiently as wild‐type FVa. FVa inactivation assays suggested that the mutation impairs its affinity for phospholipid membranes and with protein S within the complex. Conclusions FVa plays a central role in the formation of its inactivation complex. Furthermore, membrane proximal interactions among FVa, APC, and protein S are essential for its cofactor function.