Targeting of signal sequenceless proteins for export in Escherichia coli with altered protein translocase.
Most extracytoplasmic proteins are synthesized with an N-terminal signal sequence that targets them to the export apparatus. Escherichia coli prlA mutants (altered in the secY gene) are able to export cell envelope proteins lacking any signal sequence. In order to understand how such proteins are targeted for export, we isolated mutations in a signal sequenceless version of alkaline phosphatase that block its export in a prlA mutant. The mutations introduce basic amino acyl residues near the N-terminus of alkaline phosphatase. These changes do not disrupt an N-terminal export signal in this protein since the first 25 amino acids can be removed without affecting its export competence. These findings suggest that signal sequenceless alkaline phosphatase does not contain a discrete domain that targets it for export and may be targeted simply because it remains unfolded in the cytoplasm. We propose that basic amino acids near the N-terminus of a signal sequenceless protein affect its insertion into the translocation apparatus after it has been targeted for export. These findings allow the formulation of a model for the entry of proteins into the membrane-embedded export machinery.
Department of Microbiology and Molecular Genetics, Harvard Medical School, Boston, MA 02115, USA.
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Prinz
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Spiess
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Ehrmann
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Schierle
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Beckwith
D000469:Alkaline Phosphatase / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D000681:Amylases / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics*; D001692:Biological Transport; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins; D009154:Mutation; D021382:Protein Sorting Signals / Q000502:physiology*; D000069816:SEC Translocation Channels; D013489:Suppression, Genetic
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D021382:Protein Sorting Signals; D000069816:SEC Translocation Channels; C057841:SecY protein, E coli; D000469:Alkaline Phosphatase; D000681:Amylases