TABLE 3.
Characterization of constitutive SSY5 mutants
| SSY5 allele | Amino acid substitution(s) | AGP1 activation (Miller units)a | % Basal Stp1p processingb (SEM) | EC50c (μM [SEM]) |
|---|---|---|---|---|
| SSY5 wild type | None | 0.121 | 5.5 (1.2) | 8.1 (1.9) |
| SSY5-1 | E131K, L288V | 1.461 | ND | ND |
| SSY5-2 | S430N, F575V, L645M | 1.916 | ND | ND |
| SSY5-3 | V412A, K581N, P632H | 1.297 | ND | ND |
| SSY5-10 | K304R, Q576P | 2.187 | ND | ND |
| SSY5-12 | K268M, G562C, F575V | 2.158 | ND | ND |
| SSY5-13 | E131K | 1.015 | 15.5 (2.1) | 10.3 (2.2) |
| SSY5-14 | F575V | 1.201 | 29.5 (5.5) | 1.4 (0.2) |
| SSY5-15 | Q576P | 0.810 | 32.1 (2.7) | 0.8 (0.2) |
| SSY5-16 | K581N | 0.145 | ND | ND |
| SSY5-17 | P632H | 0.087 | ND | ND |
| SSY5-18 | K581N, P632H | 0.730 | 9.3 (1.2) | 2.1 (0.4) |
| SSY5-6d | E512K | 1.646 | 36.9 (3.6) | 3.2 (0.6) |
a
β-Galactosidase activity in M5361 (ssy5Δ AGP1::PAGP1-lacZ-kanMX) cells transformed with the centromere-based vector pRS316 carrying the SSY5 mutant gene and grown in SD medium without amino acids.
b
Stp1p processing in M5444 (ssy5Δ STP1::ZZ-kanMX) cells transformed with the centromeric vector pRS316 carrying the SSY5 mutant gene and grown in SD medium without amino acids. The average of at least two individual determinations is shown. ND, not determined.
c
EC50 values (apparent Kd) were determined as described (27, 28). The averages of at least two individual determinations are shown.
d
Data are from reference 28.