Encoded by the THBD gene, thrombomodulin is also known as CD141 antigen. The deduced amino acid sequence of human THBD (Accession # NP_000352) predicts a signal peptide (aa 1-18) and a mature chain (aa 19-575) that consists of following domains: C-type lectin (aa 35-169), EGF-like (aa 242-281, aa 284-324, aa 325-363, aa 365-405, aa 404-440, and aa 441-481), Ser/Thr-rich (aa 490-506), transmembrane (aa 516 to 539) and cytoplasmic (aa 540-575) (1). rhTHBD consists of aa 19-515, corresponding to the extracellular portion of the type I membrane protein. Predominantly synthesized by vascular endothelial cells, THBD inhibits coagulation and fibrinolysis (1-3). It functions as a cell surface receptor and an essential cofactor for active thrombin, which in turn activates protein C and thrombin-activatable fibrinolysis inhibitor (TAFI), also known as carboxypeptidase B2 (CPB2). Activated protein C (APC), facilitated by protein S, degrades coagulation factors Va and VIIIa, which are required for thrombin activation. Activated CPB2 cleaves basic C-terminal amino acid residues of its substrates, including fibrin, preventing the conversion of plasminogen to plasmin. In addition, THBD gene polymorphisims are associated with human disease and THBD plays a role in thrombosis, stroke, arteriosclerosis, and cancer (4). For example, increased serum levels of THBD, due to protease cleavage, have been associated with smoking, cardiac surgery, atherosclerosis, liver cirrhosis, diabetes mellitus, cerebral and myocardial infarction, and multiple sclerosis (5).
Source: Recombinant Human Thrombomodulin encoded by the THBD gene (rhTHBD; residues 19-515; GenBank Accession # NP_000352) was expressed with a N-terminal human CD33 signal peptide and a C-terminal 6-His tag in a murine myeloma cell line, NS0.
Molecular Mass: The purified, secreted rhTHBD has a N-terminal sequence of A19PAEPQPGGS. The 503 amino acid residue rhTHBD predicts a molecular mass of 53 kD and migrates with an apparent molecular mass of 95 kD in SDS-PAGE under reducing conditions.
Endotoxin Level: < 1.0 EU per 1 μg of the protein as determined by the LAL method.
Activity: Measured by its ability to mediate thrombin activation of protein C. See Activity Assay Protocol for details. The specific activity of rhProtein C, measured with 5 ng of rhThrombin, 5 ng of rhTHBD, 100 ng of rhProtein C, and 0.1 mM Boc-D(OBzl)PR-AMC in 100 μL of volume at room temperature, is > 300 pmoles/min/μg.
Storage: This protein can be aliquoted and stored under sterile conditions at -20°C in a manual defrost freezer for up to six months without a significant loss of activity.
