Translocation of a folded protein across the outer membrane in Escherichia coli.

A mutation in the Escherichia coli dsbA gene (coding for a periplasmic disulfide oxidoreductase) reduces the rate of disulfide bond formation in the enzyme pullulanase and also reduces the rate at which the enzyme is secreted to the cell surface, as measured by protease accessibility. The enzyme did not become protease accessible when disulfide bond formation was completely prevented in the mutant strain by carboxymethylation. These results indicate that a disulfide bond may be required for, and certainly does not impede, the translocation of pullulanase across the outer membrane. Since it is unlikely that a disulfide bond could be formed and then reduced again in the periplasm, these results would appear to strengthen the argument that pullulanase polypeptides fold into or close to their final conformation before they are transported across the outer membrane. It is suggested that this might be a feature common to all proteins that are secreted by other Gram-negative bacteria by a pullulanase-like pathway.