Enzymatic properties of the Arg67His mutant thrombin toward synthetic and natural substrates/ligands
| . | WT . | Arg67His . | . | ||
|---|---|---|---|---|---|
| Enzymatic properties | |||||
| Substrate | kcat(s−1) | Km(μM) | kcat(s−1) | Km(μM) | ΔΔG* (Kcal/mol) |
| Phe-Pip-Arg-pNA | 85 ± 9 | 2.1 ± 0.2 | 81 ± 8 | 2.0 ± 0.1 | 0 |
| Fibrinogen (Aα) | 64.3 ± 1.3 | 2.9 ± 0.2 | 32.5 ± 2.9 | 37.8 ± 6.5 | 1.9 |
| TR 38-60 peptide | 106 ± 2.3 | 2.7 ± 0.2 | 49.9 ± 3.2 | 62.2 ± 8.3 | 2.6 |
| kcat/Km (M−1s−1) | kcat/Km(M−1 s−1) | ||||
| PC (no TM) | 5.03 ± 0.4 × 103 | 4.7 ± 0.2 × 103 | 0 | ||
| PC (with 100 nM TM) | 2.3 × 106 | 3.7 × 105 | 1.1 | ||
| . | WT . | Arg67His . | . | ||
|---|---|---|---|---|---|
| Enzymatic properties | |||||
| Substrate | kcat(s−1) | Km(μM) | kcat(s−1) | Km(μM) | ΔΔG* (Kcal/mol) |
| Phe-Pip-Arg-pNA | 85 ± 9 | 2.1 ± 0.2 | 81 ± 8 | 2.0 ± 0.1 | 0 |
| Fibrinogen (Aα) | 64.3 ± 1.3 | 2.9 ± 0.2 | 32.5 ± 2.9 | 37.8 ± 6.5 | 1.9 |
| TR 38-60 peptide | 106 ± 2.3 | 2.7 ± 0.2 | 49.9 ± 3.2 | 62.2 ± 8.3 | 2.6 |
| kcat/Km (M−1s−1) | kcat/Km(M−1 s−1) | ||||
| PC (no TM) | 5.03 ± 0.4 × 103 | 4.7 ± 0.2 × 103 | 0 | ||
| PC (with 100 nM TM) | 2.3 × 106 | 3.7 × 105 | 1.1 | ||
ΔΔG is the difference in free energy of activation for substrate hydrolysis, calculated as RT ln (Rwt/Rm) where Rwt and Rm are the kcat/Km pertaining to substrate hydrolysis by WT and Arg67His mutant, respectively. R is the gas constant and T the absolute temperature.