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. 2014 Mar 27;3(4):297–306. doi: 10.1242/bio.20146866

Fig. 5. Whole-cell MTSEA-labeling of receptors containing cysteine at position 106.

Fig. 5.

Cells were labeled with MTSEA in the absence (−) or presence (+) of α-factor. Detergent solubilized receptors that had been labeled with MTSEA were collected on avidin beads and analyzed by SDS-PAGE and western blotting with anti-HA antiserum. (A) Strains were DJ213-7-3 containing plasmid pDJ683 (Ste2-Y106C-HA) and strain DJ213-7-3 containing plasmid pDJ684 (Ste2-Y106C,F204S-HA). (B) Plasmids encoding Ste2-Y106C-HA or Ste2-Y106C, F204S-HA were introduced into the strains expressing wild type or Ste2-F204S under the direction of the GPD transcriptional promoter. Strains were DJ484-1 containing plasmid pDJ683 (Ste2/Ste2-Y106C-HA), DJ484-1 containing plasmid pDJ684 (Ste2/Ste2-Y106C, F204S-HA), DJ482-1 containing plasmid pDJ683 (Ste2-F204S/Ste2-Y106C-HA), and DJ482-1 containing plasmid pDJ684 (Ste2-F204S/Ste2-Y106C,F204S-HA). For the total protein control (lower panel), the detergent solubilized receptors were analyzed before the avidin purification step. Immunoblots are developed with anti-HA antiserum. Molecular weights of markers are indicated in kDa at the left.