Table 1.
Steady-state kinetic parameters of selected tHisA variants compared with wild-type enzymes tHisA and tTrpF
| Enzyme | TrpF reaction
|
HisA reaction
|
||||
|---|---|---|---|---|---|---|
| kcat, s−1 | KmPRA, mM | kcat/KmPRA, mM−1⋅s−1 | kcat, s−1 | KmProFAR, μM | kcat/KmProFAR, μM−1⋅s−1 | |
| tHisA | 0 | ND | ND | 0.70 | 0.60 | 1.2 |
| tHisA_1* | >0.046 | >1.0 | 0.046 | 0.061 | 1.32 | 0.046 |
| tHisA_2* | >0.049 | >0.1 | 0.49 | ≈2⋅10−4 | ND | ND |
| tHisA_Asp127 Val* | >0.012 | >0.1 | 0.12 | ≈1⋅10−4 | ND | ND |
| tTrpF† | 3.7 | 2.8⋅10−4 | 13.3⋅103 | 0 | ND | ND |
Buffer conditions: 50 mM Tris⋅HCl, pH 7.5 at 25°C, containing 4 mM MgEDTA and 2 mM DTT (TrpF reaction), or 2 mM NaEDTA and 1 mM DTT (HisA reaction). ND, because of the very low kcat value, Km and therefore also kcat/Km could not be determined.
*
Only minimal values for kcat and KmPRA could be determined, because KmPRA is at least 10 times as high as the maximally applied substrate concentrations.
†
Data from ref. 21.