Protein S1₁₀₆ affects <i>E. coli</i> growth by displacing native S1 from the ribosome.

<p>(<b>A</b>) In contrast to synthesis of protein S1_87–194 (representing domain D2, -•-), synthesis of S1₁₀₆ and S1₁₉₄ (representing domains D1 (-▪-) and D1-2 (-▴-)) inhibits bacterial growth. <i>E. coli</i> strain JE28 harbouring plasmids pProEX-HTb (-◊-), pPro-S1D1F (-▪-), pPro-S1D2F (-•-) and pPro-S1D1-2F (-▴-) were grown in LB medium containing ampicilin (100 µg/ml) and kanamycin (20 µg/ml). At OD₆₀₀ of 0.2–0.25 (indicated by an arrow) 50 µM IPTG was added to the cultures. Aliquots were withdrawn from each culture for ribosome preparation 1 hour upon induction. (<b>B</b>) Proteins present in S30 extracts (lanes 1, 3, 5, and 7) and 70 S ribosomes (lanes 2, 4, 6, and 8) prepared from cells without overexpression (lanes 1 and 2), and cells overexpressing S1₁₀₆ (lanes 3 and 4), S1_87–194 (lanes 5 and 6), or S1₁₉₄ (Lanes 7 and 8) were separated on a 12.5% SDS-PAGE and presence of protein S1 and its variants on 70 S ribosomes was checked by western blot analysis using anti-S1 antibodies (panel a), anti-FLAG (panel b) and anti-L2 antibodies (panel c), which served as loading control. The positions of the respective proteins are indicated to the right. (<b>C</b>) The N-terminal domain of S1 is required for assembly to the ribosome. Equimolar amounts of HIS-tagged ribosomes (lanes 1 and 3) and ribosome free S100 extract (lanes 2 and 4) purified from <i>E. coli</i> strain JE28 overexpressing FLAG-tagged proteins S1 (lanes 1 and 2) and S1_87–557, lacking domain D1 (lanes 3 and 4) were separated on a 12.5% SDS-PAGE. The presence of S1 and S1_87–557 was determined by western blot analysis employing anti-FLAG antibodies (panel a) and anti-L2 antibodies (panel b), which served as loading control.