The stable BRP signal peptide causes lethality but is unable to provoke the translocation of cloacin DF13 across the cytoplasmic membrane of Escherichia coli.
The bacteriocin release protein (BRP) mediates the secretion of cloacin DF13. The BRP precursor is slowly processed to yield the mature BRP and its stable signal peptide which is also involved in cloacin DF13 secretion. The function of the stable BRP signal peptide was analysed by constructing two plasmids. First, the stable BRP signal peptide was fused to the murein lipoprotein and, second, a stop codon was introduced after the BRP signal sequence. Exchange of the unstable murein lipoprotein signal peptide for the stable BRP signal peptide resulted in an accumulation of precursors of the hybrid murein lipoprotein. This indicated that the BRP signal peptide, as part of this hybrid precursor, is responsible for the slow processing. The stable BRP signal peptide itself was not able to direct the transfer of cloacin DF13 into the periplasmic space or into the culture medium. Over-expression of the BRP signal peptide was lethal and caused 'lysis'. Subcellular fractionation experiments revealed that the BRP signal peptide is located exclusively in the cytoplasmic membrane whereas the mature BRP, targeted by either the stable BRP signal peptide or the unstable Lpp signal peptide, is located in both the cytoplasmic and outer membrane. These results are in agreement with the hypothesis that the stable signal peptide and the mature BRP together are required for the passage of cloacin DF13 across the cell envelope.
van der Wal
F J
FJ
Department of Molecular Microbiology, Faculty of Biology, Vrije Universiteit, Amsterdam, The Netherlands.
Oudega
B
B
Kater
M M
MM
ten Hagen-Jongman
C M
CM
de Graaf
F K
FK
Luirink
J
J
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001430:Bacteriocins; D001483:Base Sequence; D001692:Biological Transport; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D029968:Escherichia coli Proteins; D008969:Molecular Sequence Data; D010957:Plasmids; D011499:Protein Processing, Post-Translational; D021382:Protein Sorting Signals / Q000235:genetics / Q000378:metabolism*; D001618:beta-Lactamases / Q000235:genetics
D001426:Bacterial Proteins; D001430:Bacteriocins; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D021382:Protein Sorting Signals; C027355:cloacin DF13 protein, E coli; C052368:cloacin DF13-encoded bacteriocin release protein, E coli; D001618:beta-Lactamases