Reactivity | HuSpecies Glossary |
Applications | Enzyme Activity |
Format | Carrier-Free |
Details of Functionality | Measured by its ability to cleave the fluorogenic peptide substrate Boc-VPR-AMC (Catalog # ES011). The specific activity is >20,000 pmol/min/µg, as measured under the described conditions. |
Source | Mouse myeloma cell line, NS0-derived human Coagulation Factor II/Thrombin protein Met1-Glu622 with a C-terminal 10-His tag The proenzyme was purified, activated and further purified. |
Accession # | |
N-terminal Sequence | Thr328, Ile364 & Val515 |
Structure / Form | Active form |
Protein/Peptide Type | Recombinant Enzymes |
Gene | F2 |
Purity | >95%, by SDS-PAGE under reducing conditions and visualized by silver stain |
Endotoxin Note | <1.0 EU per 1 μg of the protein by the LAL method. |
Dilutions |
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Theoretical MW | 4 kDa (light chain A), 31 kDa (heavy chain B), 14 kDa (C-terminal fragment). Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors. |
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SDS-PAGE | 12 kDa, 21 kDa and 23 kDa, reducing conditions |
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Publications |
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Storage | Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
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Buffer | Lyophilized from a 0.2 μm filtered solution in HEPES and NaCl. |
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Purity | >95%, by SDS-PAGE under reducing conditions and visualized by silver stain |
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Reconstitution Instructions | Reconstitute at 100 μg/mL in sterile 50 mM HEPES and 580 mM NaCl, pH 7.5. |
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Assay Procedure |
*Adjusted for Substrate Blank **Derived using calibration standard 7-amino, 4-Methyl Coumarin (Sigma, Catalog # A-9891). Per Well:
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Coagulation Factor II, commonly known as thrombin, is an essential component of the coagulation cascade in which it converts fibrinogen to fibrin, activates factors V, VII, VIII, XIII and forms complexes with protein C and thrombomodulin (1). It also activates platelets and regulates the behavior of additional cells through protease‑activated receptors (PARs) (2). It may have either protective or deleterious functions, depending on the level and location (3). Its activity is regulated by endogenous inhibitors such as anti-thrombin III (serpin C1) or heparin cofactor II (serpin D1). A plasma serine protease, thrombin is synthesized in the liver as a 622 amino acid precursor with a 24 amino acid signal peptide. Cleavage by itself or by similar enzymes converts the proenzyme to three forms designated as alpha ‑, beta ‑ and gamma ‑thrombin. Composed of a disulfide bond-linked dimer of the light chain (A) (residues 328-363) and the heavy chain (B) (residues 364‑622), alpha -thrombin displays the diverse functions as described above. In comparison, the further processed B chains of beta - and gamma -thrombin have no known physiological function, but retain most of the activity towards small synthetic substrates (4). The recombinant human thrombin may be used to process proteins containing the thrombin cleavage site. The conditions for processing different proteins, such as the ratio of protein/thrombin, buffer components, temperature and time of incubation, may vary and should be empirically determined.
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