Macromolecular import into Escherichia coli: the TolA C-terminal domain changes conformation when interacting with the colicin A toxin.
Various macromolecules such as bacteriotoxins and phage DNA parasitize some envelope proteins of Escherichia coli to infect the bacteria. A two-step import mechanism involves the primary interaction with an outer membrane receptor or with a pilus followed by the translocation across the outer membrane. However, this second step is poorly understood. It was shown that the TolA, TolQ, and TolR proteins play a critical role in the translocation of group A colicins and filamentous bacteriophage minor coat proteins (g3p). Translocation of these proteins requires the interaction of their N-terminal domain with the C-terminal domain of TolA (TolAIII). In this work, short soluble TolAIII domains were overproduced in the cytoplasm and in the periplasm of E. coli. In TolAIII, the two cysteine residues were found to be reduced in the cytoplasmic form and oxidized in the periplasmic form. The interaction of TolAIII with the N-terminal domain of colicin A (ATh) is observed in the presence and in the absence of the disulfide bridge. The complex formation of TolAIII and ATh was found to be independent of the ionic strength. An NMR study of TolAIII, both free and bound, shows a significant structural change when interacting with ATh, in the presence or absence of the disulfide bridge. In contrast, such a structural modification was not observed when TolAIII interacts with g3p N1. These results suggest that bacteriotoxins and Ff bacteriophages parasitize E. coli using different interactions between TolA and the translocation domain of the colicin and g3p protein, respectively.
Deprez
Christophe
C
Institut de Biologie Structurale Jean-Pierre Ebel, CEA-CNRS-UJF, UMR 5075, 41 rue Jules Horowitz, 38027 Grenoble cedex 1, France.
Blanchard
Laurence
L
Guerlesquin
Françoise
F
Gavioli
Marthe
M
Simorre
Jean-Pierre
JP
Lazdunski
Claude
C
Marion
Dominique
D
Lloubès
Roland
R
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D001483:Base Sequence; D001692:Biological Transport; D002942:Circular Dichroism; D003087:Colicins / Q000378:metabolism*; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D011487:Protein Conformation; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D001426:Bacterial Proteins; D003087:Colicins; D017931:DNA Primers; D029968:Escherichia coli Proteins; C062093:tolA protein, E coli