Basigin-MCT1 fusion constructs, expression in yeast, and transport functionality.

(A) Shown are fusion constructs of basigin (black/gray; PDB #6LZ0) with MCT1 (orange; PDB #6LZ0). Disulfide bridges are shown as orange spheres, and Glu218 of the basigin transmembrane helix is highlighted in red. (B) Western blot showing expression of MCT1 alone (58 kDa), and fused with BSGΔIg (WT, 67 kDa) or the E27R point mutant. Proteins were detected using a Penta-His antibody. (C) Live-cell confocal microscopy of fusion proteins via C-terminal GFP and soluble GFP (—). (D, E) Uptake of ¹⁴C-labeled l-lactate into jen1Δ ady2Δ yeast at pH 6.8 and a 1 mM inward gradient. Shown are curves for non-expressing cells (background, D, ○), cells expressing MCT1 alone (D, ●), and in fusion with BSGΔIg (E, ■), or BSGΔIg E27R (E, □). The data are normalized to 1 mg of cells and the background of non-expressing cells was subtracted. Error bars indicate ± S.E.M. from three biological replicates.