FIG 4.

SecA interacts with nascent chains when they reach a length of >116 amino acids. (A) Diagram of the arrested SecM-PhoA fusions. The signal sequence, the 18-amino-acid (aa) amino arrest sequence, and the deleted regions are indicated. Numbering refers to the number of amino acids from the start codon to the arrest-inducing proline in the SecM arrest sequence. (B) SecA (0.5 μM) was incubated with the indicated ribosome-nascent chain complex (RNC; 0.5 μM) in the presence of 250 mM potassium acetate. Ribosomes were subsequently pelleted through a 30% sucrose cushion at >200,000 × g, and the pellet fractions were analyzed by SDS-PAGE and Coomassie staining. The running positions of SecA and the ribosomal proteins are indicated. Quantitation of the SecA signal relative to the S1 signal in the Coomassie-stained gel for each RNC is indicated below.