Table 2.
Molar Cd/GS ratios of LMW-Cd·GS and HMW-Cd·GS complexes fractionated by Sephadex G-15 and Mono-Q chromatography (Fig. 2) before and after MgATP-dependent, uncoupler-insensitive uptake by vacuolar membrane vesicles purified from DTY165 and DTY167 cells
| Fraction |
109Cd uptake, nmol/mg/10 min
|
Molar ratio Cd/GS
|
||
|---|---|---|---|---|
| DTY165 | DTY167 | Before uptake | After uptake | |
| Sephadex G-15 | ||||
| HMW-Cd·GS | 0.44 ± 0.09 | |||
| LMW-Cd·GS | 0.67 ± 0.04 | |||
| Mono-Q | ||||
| HMW-Cd·GS | 66.3 ± 2.7 | 5.6 ± 2.6 | 0.49 ± 0.17 | 0.49 ± 0.01 |
| LMW-Cd·GS | 4.4 ± 0.8 | 3.9 ± 1.4 | 0.86 ± 0.07 | |
| After 2-ME | ||||
| HMW-Cd·GS | 11.9 ± 2.4 | 4.4 ± 3.0 | ||
Cd/GS ratios were estimated from the 109Cd/[3H] radioisotope ratios of samples prepared from 109CdSO4 and [3H]GSH. HMW-109Cd·[3H]GS was pretreated with 2-mercaptoethanol (2-ME) by heating a 1:4 mixture of HMW-109Cd.[3H]GS with 2-ME at 60°C for 10 min before measuring 109Cd2+ uptake. Uptake was measured using 50 μM concentrations (as Cd) of the complexes indicated in standard uptake medium containing 5 μM gramicidin-D. Values shown are means ± SE (n = 3–6).