Table 5. The maximum emission wavelength (λmax) and intrinsic fluorescence intensity at λmax of tryptophan fluorescence of adGSTD4-4 and the engineered enzymes.
The excitation wavelength (λex) was set at 295 nm, and emission was scanned from 300–450 nm. Samples (n=3) contained 0.1 mg/ml protein in 0.1 M potassium phosphate buffer, pH 6.5. The percentage intensities compared with wild-type enzyme were measured at fluorescence λmax averaged over three scans, corrected for dilution and inner-filter effects.
| Intrinsic fluorescence (λex 295 nm) | ||
|---|---|---|
| Enzyme | λmax (nm) | % intensity |
| Wild-type | 333±0 | 100 |
| L33A | 334±0 | 60.8* |
| L33Y | 334±0 | 51.6* |
| L33F | 334±0 | 47.9* |
| L33I | 333±0 | 109.1 |
| L6A | 334±0 | 138.6* |
| T31A | 334±0 | 99.1 |
| I52A | 339±0 | 98.8 |
| I52L | 333±0 | 115.5* |
| E37A | 333±0 | 66.8* |
| E37Q | 334±0 | 107.7 |
| K40A | 334±0 | 68.4* |
| E42A | 333±0 | 75.5* |
| A35R | 334±0 | 102.1 |
ANOVA analysis revealed a significant difference compared with the wild-type enzyme, where indicated (*P<0.001); the absence of an asterisk indicates no significant difference compared with the wild-type.