Fig. 5. Sensitization of SecE to FtsH by addition of extra amino acids to its N-terminus. (A) Amino acid sequences at the N-terminal cytosolic regions of SecE, HA12-SecE and Z42-SecE. The extra sequences are underlined. (B) Stability of SecE and HA12-SecE. Each of strains AR5087 (ftsH⁺; solid symbols) and AR5090 (DftsH; open symbols) was transformed either with pCM134 (SecE; circles) or pHAsecE (HA12-SecE; triangles). Plasmid-bearing cells were grown at 37°C, and induced for 10 min for lac or ara transcription. Cells were then pulse-labeled with [³⁵S]methionine for 0.5 min followed by chase with unlabeled methionine for 0, 0.5, 1 and 2 min. Labeled proteins were immunoprecipitated with anti-SecE and separated by SDS–PAGE. Radioactivities are reported as values relative to that at the 0 min chase point. (C) Stability of SecE and Z42-SecE. Cells of strains AR5087 (ftsH⁺; lanes 1–4) and AR5090 (ΔftsH; lanes 5–8), each carrying pKY271 (Z42-SecE/SecE), were grown, induced for lac transcription, and pulse-labeled for 0.5 min with [³⁵S]methionine followed by chase for 0 (lanes 1 and 5), 0.5 (lanes 2 and 6), 1 (lanes 3 and 7) and 2 min (lanes 4 and 8). Labeled proteins were immunoprecipitated with anti-SecE.