Table 1. Functional properties of various mCST constructs.
The Kd for the indicated ligand was determined by either: 1) a full SPA titration as shown in Figure 1A 2) fitting the two-point SPA data from Figure 5H to get a crude estimation of the Kd, or 3) fitting the tryptophan fluorescence quenching data shown in Figure 5G (Kd’s were not fit for the Trp207 mutants which showed no quenching). The Km and Vmax were determined from data shown in Figure 1C.
| Construct | Ligand | Kd (µM) | Km (µM) | Vmax (nmol/mol mCST/min) | ||
|---|---|---|---|---|---|---|
| SPA titration | SPA 2-point | Trp-FL | ||||
| mCST | CMP | 6.3 ± 1.2 | 5.5 ± 1.6 | 2.9 ± 1.1 | ||
| CMP-Sia | 482 ± 147 | 58.1 ± 13.2 | 6.5 ± 0.6 | |||
| mCST∆C | CMP | 1.8 ± 0.4 | 11.1 ± 2.9 | |||
| CMP-Sia | 295 ± 100 | 62.0 ± 14.7 | 5.3 ± 0.5 | |||
| UMP | 1621 ± 1119 | |||||
| GMP | 5376 ± 1151 | |||||
| mCST (W145L) | CMP | 0.8 ± 0.3 | 3.3 ± 1.0 | |||
| mCST (W160L) | CMP | 1.6 ± 0.4 | 11.9 ± 2.4 | |||
| mCST (W207L) | CMP | 1.5 ± 0.4 | N/D | |||
| mCST (W207F) | CMP | 3.8 ± 1.5 | N/D | |||
| mCST (W247L) | CMP | 2.4 ± 1.5 | 7.4 ± 2.6 | |||