Interactions between peptidoglycan and the ExeAB complex during assembly of the type II secretin of Aeromonas hydrophila.
Aeromonas hydrophila transports extracellular protein toxins via the type II secretion system, an export mechanism comprised of numerous proteins that spans both the inner and outer membranes. Two components of this secretion system, ExeA and ExeB, form a complex in the inner membrane that functions to locate and/or assemble the ExeD secretin in the outer membrane. In the studies reported here, two-codon insertion mutagenesis of exeA revealed that an insertion at amino acid 495 in the C-terminal region of ExeA did not alter ExeAB complex formation yet completely abrogated its involvement in ExeD secretin assembly and thus rendered the bacteria secretion negative. In silico analysis of protein motifs with similar amino acid profiles revealed that this amino acid is located within a putative peptidoglycan (PG) binding motif in the periplasmic domain of ExeA. Substitution mutations of three highly conserved amino acids in the motif were constructed. In cells expressing each of these mutants, the ability to assemble the ExeD secretin or secrete aerolysin was lost, while ExeA retained the ability to form a complex with ExeB. In in vivo cross-linking experiments, wild-type ExeA could be cross-linked to PG, whereas the three substitution mutants of ExeA could not. These data indicate that PG binding and/or remodelling plays a role in the function of the ExeAB complex during assembly of the ExeD secretin.
Howard
S Peter
SP
Department of Microbiology and Immunology, University of Saskatchewan, Saskatoon, Canada S7N 5E5. peter.howard@usask.ca
Gebhart
Carol
C
Langen
Geoffrey R
GR
Li
Gang
G
Strozen
Timothy G
TG
D016980:Aeromonas hydrophila / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001427:Bacterial Toxins / Q000378:metabolism; D001665:Binding Sites; D003062:Codon / Q000235:genetics; D008565:Membrane Proteins / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D010457:Peptidoglycan / Q000378:metabolism*; D052899:Pore Forming Cytotoxic Proteins; D017434:Protein Structure, Tertiary; D021381:Protein Transport
D001426:Bacterial Proteins; D001427:Bacterial Toxins; D003062:Codon; C090402:ExeA protein, Aeromonas hydrophila; C090403:ExeB protein, Aeromonas hydrophila; C094204:ExeD protein, Aeromonas; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D010457:Peptidoglycan; D052899:Pore Forming Cytotoxic Proteins; C032676:aerolysin