Identification of amino acid residues essential for the enzymatic activities of pertussis toxin.
The enzymatic ADP-ribosyltransferase activity associated with the S1 subunit of pertussis toxin is considered to be responsible for its biological effects. Although pertussis toxin has no significant homology to other ADP-ribosylating toxins such as diphtheria toxin and Pseudomonas aeruginosa exotoxin A, the results presented in this paper show that, as for diphtheria toxin and exotoxin A, tryptophan and glutamic acid residues are essential for the enzymatic activities of pertussis toxin. Moreover, a structural motif can be identified around the critical glutamic acid residue. Chemical modification or site-directed deletion or replacement of Trp-26 abolishes ADP-ribosyltransferase and the associated NAD glycohydrolase activities. Both enzymatic activities are also abolished when Glu-129 is deleted or replaced by aspartic acid. Mutations at the Glu-106 position do not significantly reduce the enzymatic activities of the S1 subunit. The mutations do not affect the ability of the different S1 forms to be recognized by a variety of monoclonal antibodies, including neutralizing antibodies. Pertussis toxin containing a deletion or replacement of Trp-26, Glu-129, or both in the S1 subunit should thus be devoid of toxic activities without losing its reactivity with protective antibodies and, therefore, could be safely included in new generation vaccines against whooping cough.
Locht
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Department of Molecular and Cellular Biology, Smith Kline-Biologicals, Rixensart, Belgium.
Capiau
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Feron
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D015077:2-Hydroxy-5-nitrobenzyl Bromide / Q000494:pharmacology; D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D000595:Amino Acid Sequence; D000596:Amino Acids; D001435:Bacteriophages / Q000235:genetics; D001483:Base Sequence; D055598:Chemical Phenomena; D002621:Chemistry; D004274:DNA, Recombinant; D000939:Epitopes / Q000276:immunology; D004926:Escherichia coli / Q000235:genetics; D019204:GTP-Binding Proteins / Q000378:metabolism; D005786:Gene Expression Regulation; D005971:Glutamates; D018698:Glutamic Acid; D008969:Molecular Sequence Data; D009154:Mutation; D009244:NAD+ Nucleosidase / Q000378:metabolism; D037342:Pertussis Toxin; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D013329:Structure-Activity Relationship; D014162:Transfection; D014364:Tryptophan; D010566:Virulence Factors, Bordetella / Q000235:genetics / Q000276:immunology / Q000378:metabolism*
D000596:Amino Acids; D004274:DNA, Recombinant; D000939:Epitopes; D005971:Glutamates; D010566:Virulence Factors, Bordetella; D000246:Adenosine Diphosphate Ribose; D018698:Glutamic Acid; D015077:2-Hydroxy-5-nitrobenzyl Bromide; D014364:Tryptophan; D011065:Poly(ADP-ribose) Polymerases; D037342:Pertussis Toxin; D009244:NAD+ Nucleosidase; D019204:GTP-Binding Proteins