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Addition of the nonmetabolizable amino acid cysteine or glutamate analog γ-MGA results in less Gap1p localized at the plasma membrane. A wild-type PADH1-GAP1-HA strain (CKY759) was grown on minimal ammonia medium to exponential phase at 24°C. Glutamate, cysteine, or γ-MGA was added to 0.75 mM and, after 2 h, cells were harvested for extract preparation and fractionation on 20–60% sucrose density gradients.
