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The catalytic cycle of the Escherichia coli SecA ATPase comprises two distinct preprotein translocation events
Author(s) -
van der Wolk Jeroen P.W.,
de Wit Janny G.,
Driessen Arnold J.M.
Publication year - 1997
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1093/emboj/16.24.7297
Subject(s) - biology , escherichia coli , chromosomal translocation , escherichia coli proteins , atpase , microbiology and biotechnology , biochemistry , enzyme , gene
SecA is the ATP‐dependent force generator in the Escherichia coli precursor protein translocation cascade, and is bound at the membrane surface to the integral membrane domain of the preprotein translocase. Preproteins are thought to be translocated in a stepwise manner by nucleotide‐dependent cycles of SecA membrane insertion and de‐insertion, or as large polypeptide segments by the protonmotive force (Δp) in the absence of SecA. To determine the step size of a complete ATP‐ and SecA‐dependent catalytic cycle, translocation intermediates of the preprotein proOmpA were generated at limiting SecA translocation ATPase activity. Distinct intermediates were formed, spaced by intervals of ∼5 kDa. Inhibition of the SecA ATPase by azide trapped SecA in a membrane‐inserted state and shifted the step size to 2–2.5 kDa. The latter corresponds to the translocation elicited by binding of non‐hydrolysable ATP analogues to SecA, or by the re‐binding of partially translocated polypeptide chains by SecA. Therefore, a complete catalytic cycle of the preprotein translocase permits the stepwise translocation of 5 kDa polypeptide segments by two consecutive events, i.e. ∼2.5 kDa upon binding of the polypeptide by SecA, and another 2.5 kDa upon binding of ATP to SecA.