Table 3.
Transient expression of novel SMPD1 missense mutations in 293-T cells.
| Construct/mutation | Mean ASM activity, U/mg protein | % Wild-type activity | Stability,a % initial activity | Predicted phenotype |
|---|---|---|---|---|
| ASM (wild type) | 546 | 100 | 55.9 | WT |
| W211R | 2.34 | 0.43 | NDb | A |
| D253H | 0.13 | 0.02 | NDb | A |
| V314M | 118 | 21.7 | 18.2 | B |
| H427R | 0.10 | 0.02 | NDb | A |
| N522S | 55.1 | 10.1 | 45 | B |
| Q525H | 349 | 63.9 | 19.9 | B |
| H577R | 2.66 | 0.49 | NDb | A |
a
For stability studies, 24 h after transfection cell extracts were prepared and incubated at 55°C, pH 7.0, for 30 min prior to enzyme assay.
b
ND: thermostability was not determined for these alleles, because the expressed activities were already at the lower limit of detectability.