Characterization of the C-terminal domain essential for toxic activity of adenylate cyclase toxin.
Adenylate cyclase toxin (CyaA) of Bordetella pertussis belongs to the RTX family of toxins. These toxins are characterized by a series of glycine- and aspartaterich nonapeptide repeats located at the C-terminal half of the toxin molecules. For activity, RTX toxins require Ca2+, which is bound through the repeat region. Here, we identified a stretch of 15 amino acids (block A) that is located C-terminally to the repeat and is essential for the toxic activity of CyaA. Block A is required for the insertion of CyaA into the plasma membranes of host cells. Mixing of a short polypeptide composed of block A and eight Ca2+ binding repeats with a mutant of CyaA lacking block A restores toxic activity fully. This in vitro interpolypeptide complementation is achieved only when block A is present together with the Ca2+ binding repeats on the same polypeptide. Neither a short polypeptide composed of block A only nor a polypeptide consisting of eight Ca2+ binding repeats, or a mixture of these two polypeptides, complement toxic activity. It is suggested that functional complementation occurs because of binding between the Ca2+ binding repeats of the short C-terminal polypeptide and the Ca2+ binding repeats of the CyaA mutant lacking block A.
Bejerano
M
M
Department of Clinical Microbiology, Hebrew University-Hadassah Medical School, Jerusalem, Israel.
Nisan
I
I
Ludwig
A
A
Goebel
W
W
Hanski
E
E
D037361:Adenylate Cyclase Toxin; D000262:Adenylyl Cyclases / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D001886:Bordetella pertussis / Q000235:genetics / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D002874:Chromosome Mapping; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins; D015870:Gene Expression; D005816:Genetic Complementation Test; D006460:Hemolysin Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009876:Operon; D010455:Peptides / Q000378:metabolism; D011498:Protein Precursors / Q000235:genetics / Q000378:metabolism*; D010566:Virulence Factors, Bordetella / Q000235:genetics / Q000378:metabolism*