Fig. 3.

The synthesis rate of V.SecD2 is affected not only by Na⁺ but also by the absence of V.SecDF1 or the export-inhibiting azide. (A) WT V. alginolyticus grown in M63 medium supplemented with the indicated concentrations of Na⁺ were pulse-labeled for 1 min with [³⁵S]methionine, followed by immunoprecipitation of V.SecD2. Asterisks indicate nonspecific background bands. (B) WT (lane 1 and lanes 4–9), ΔsecDF1 (lane 2), and ΔsecDF2 (lane 3) strains of V. alginolyticus grown in M63-300 mM Na⁺ medium at 30 °C were pulse-labeled for 1 min without NaN₃ treatment (lanes 1, 2, 3, 8, and 9) or at the indicated time points after addition of NaN₃. Samples of equivalent radioactivities were subjected to immunoprecipitation with antibodies against V.SecD1 (Upper) and V.SecD2 (Lower). Asterisks indicate nonspecific background bands. The numbers at the bottoms show intensities of V.SecD1 relative to that in lane 1 (Upper) and those of V.SecD2 relative to that in lane 2 (Lower). (C) To verify that azide inhibits protein export in V. alginolyticus, cells were pulse-labeled for 0.5 min with (lane 1) or without (lane 2) 1 min NaN₃ pretreatment. Processing efficiencies of MBP are shown.