Membrane deinsertion of SecA underlying proton motive force-dependent stimulation of protein translocation

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Abstract
The proton motive force (PMF) renders protein translocation across the Escherichia coli membrane highly efficient, although the underlying mechanism has not been clarified. The membrane insertion and deinsertion of SecA coupled to ATP binding and hydrolysis, respectively, are thought to drive the translocation. We report here that PMF significantly decreases the level of membrane‐inserted SecA. The prlA4 mutation of SecY, which causes efficient protein translocation in the absence of PMF, was found to reduce the membrane‐inserted SecA irrespective of the presence or absence of PMF. The PMF‐dependent decrease in the membrane‐inserted SecA caused an increase in the amount of SecA released into the extra‐membrane milieu, indicating that PMF deinserts SecA from the membrane. The PMF‐dependent deinsertion reduced the amount of SecA required for maximal translocation activity. Neither ATP hydrolysis nor exchange with external SecA was required for the PMF‐dependent deinsertion of SecA. These results indicate that the SecA deinsertion is a limiting step of protein translocation and is accelerated by PMF, efficient protein translocation thereby being caused in the presence of PMF.