Figure 4. Casitone restores the overexpression levels in GCDM-grown cells.

(A) Analysis of OppA levels in total cell lysates from L. lactis NZ9000; immunoblots were developed using an anti-OppA antibody. Cells were grown in GM17, GCDM, GCDM^PDC or GCDM with various dipeptides (left panel; amino acids indicated by their one letter code) or GCDM with altered branched-chain amino acids concentrations (1X BCAA is the standard BCAA concentration in GCDM). (B) Effect of supplementation of GCDM with casein hydrolysate (GCDM^PDC; right lanes) on (membrane) protein expression in L. lactis NZ9000; GM17- (left lanes) and GCDM-grown cells (middle lanes) are shown for comparison. Cells were induced at OD₆₀₀≈0.5 with 0.1% volume of nisin A-containing NZ9700 culture supernatant and growth was continued for 2 h. Protein levels were analyzed on immunoblots, using an anti-His tag antibody. For the left panels, names in brackets indicate the subunits of the respective ABC transporters that were co-expressed without a His-tag and consequently not detected on the immunoblot. Sav1866 and RibU are examples of proteins whose expression is not affected by the medium. (C) Schematic representation of the uptake pathways for amino acids and peptides in L. lactis.