title: TFPI cofactor function of protein S: essential role of the protein S SHBG-like domain.
abstract: Protein S is a cofactor for tissue factor pathway inhibitor (TFPI), accelerating the inhibition of activated factor X (FXa). TFPI Kunitz domain 3 residue Glu226 is essential for enhancement of TFPI by protein S. To investigate the complementary functional interaction site on protein S, we screened 44 protein S point, composite or domain swap variants spanning the whole protein S molecule for their TFPI cofactor function using a thrombin generation assay. Of these variants, two protein S/growth arrest-specific 6 chimeras, with either the whole sex hormone-binding globulin (SHBG)-like domain (Val243-Ser635; chimera III) or the SHBG laminin G-type 1 subunit (Ser283-Val459; chimera I), respectively, substituted by the corresponding domain in growth arrest-specific 6, were unable to enhance TFPI. The importance of the protein S SHBG-like domain (and its laminin G-type 1 subunit) for binding and enhancement of TFPI was confirmed in FXa inhibition assays and using surface plasmon resonance. In addition, protein S bound to C4b binding protein showed greatly reduced enhancement of TFPI-mediated inhibition of FXa compared with free protein S. We show that binding of TFPI to the protein S SHBG-like domain enables TFPI to interact optimally with FXa on a phospholipid membrane.
authors.affiliation: Centre for Haematology, Faculty of Medicine, Imperial College London, London, United Kingdom;
authors.forename: Natalia
authors.initials: N
authors.lastname: Reglińska-Matveyev
authors.affiliation: Centre for Haematology, Faculty of Medicine, Imperial College London, London, United Kingdom;
authors.forename: Helena M
authors.initials: HM
authors.lastname: Andersson
authors.affiliation: Department of Internal Medicine, Faculty of Medicine, Universidade Federal de Minas Gerais, Minas Gerais, Brazil; and.
authors.forename: Suely M
authors.initials: SM
authors.lastname: Rezende
authors.affiliation: Department of Laboratory Medicine, Faculty of Medicine, Lund University, Malmö, Sweden.
authors.forename: Björn
authors.initials: B
authors.lastname: Dahlbäck
authors.affiliation: Centre for Haematology, Faculty of Medicine, Imperial College London, London, United Kingdom;
authors.forename: James T B
authors.initials: JT
authors.lastname: Crawley
authors.affiliation: Centre for Haematology, Faculty of Medicine, Imperial College London, London, United Kingdom;
authors.forename: David A
authors.initials: DA
authors.lastname: Lane
authors.affiliation: Centre for Haematology, Faculty of Medicine, Imperial College London, London, United Kingdom;
authors.forename: Josefin
authors.initials: J
authors.lastname: Ahnström
mesh_terms: D001665:Binding Sites / Q000235:genetics; D015153:Blotting, Western; D050716:Complement C4b-Binding Protein / Q000378:metabolism; D015951:Factor Xa / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D008074:Lipoproteins / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D010743:Phospholipids / Q000378:metabolism; D011486:Protein C / Q000378:metabolism; D017293:Protein S / Q000235:genetics / Q000378:metabolism*; D012738:Sex Hormone-Binding Globulin / Q000378:metabolism; D020349:Surface Plasmon Resonance; D013917:Thrombin / Q000378:metabolism; D013925:Thromboplastin / Q000378:metabolism
chemical_list: D050716:Complement C4b-Binding Protein; D008074:Lipoproteins; D010743:Phospholipids; D011486:Protein C; D017293:Protein S; D012738:Sex Hormone-Binding Globulin; C051928:lipoprotein-associated coagulation inhibitor; D013925:Thromboplastin; D013917:Thrombin; D015951:Factor Xa
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