Table 2.
enzyme | substrate | glycosylase efficiency kcat/Kd (min−1 nM−1) | 95% conf intervals | lyase catalytic rate kcat (min−1) | 95% conf intervals |
---|---|---|---|---|---|
CacOgg | 8-oxoG·C | 0.420 | 0.378−0.456 | 0.0420 | 0.0378−0.0468 |
CacOgg | 8-oxoG·A | 0.384 | 0.276−0.576 | 0.0252 | 0.0168−1.02E+06 |
M132R | 8-oxoG·C | 1.800 | 1.620−2.040 | 0.0960 | 0.0900−0.1080 |
M132R | 8-oxoG·A | 0.108 | 0.078−0.156 | 0.0342 | 0.0252−0.0444 |
F179Y | 8-oxoG·C | 0.336 | 0.300−0.384 | 0.0234 | 0.0210−0.0258 |
F179Y | 8-oxoG·A | 0.027 | 0.018−0.033 | 0.0078 | 0.0054−0.0090 |
M132R/F179Y | 8-oxoG·C | 1.080 | 0.900−1.200 | 0.1140 | 0.0960−0.1380 |
M132R/F179Y | 8-oxoG·A | 0.021 | 0.016−0.025 | 0.0072 | 0.0048−2.64E+14 |
hOGG1 | 8-oxoG·C | 0.270 | 0.180−0.390 | 0.0234 | 0.0174−0.0582 |
The efficiency (kcat/Kd) of the glycosylase reaction and kcat of the bifunctional reaction for CacOgg and variants with the substrates 8-oxoG·C and 8-oxoG·A were obtained under single turnover conditions (0.025 nM 8-oxoG·C and 0.25, 1, or 2.5 nM enzyme for the glycosylase reaction and 2.5, 12, or 25 nM enzyme for the bifunctional reaction). Each reaction was prepared as a 70 μL pool, and 10 μL aliquots were removed at various times and added to NaOH or formamide as described in Materials and Methods.