Stimulation of the Thiol-Dependent ADP-Ribosyltransferase and NAD Glycohydrolase Activities of Bordetella Pertussis Toxin by Adenine Nucleotides, Phospholipids, and Detergents

Joel Moss, Sally J. Stanley, Paul A. Watkins, Drusilla L. Burns, Charles R. Manclark, Harvey R. Kaslow, Erik L. Hewlett

Research output: Contribution to journalArticlepeer-review

Abstract

Pertussis toxin catalyzed ADP-ribosylation of the guanyl nucleotide binding protein transducin was stimulated by adenine nucleotide and either phospholipids or detergents. To determine the sites of action of these agents, their effects were examined on the transducin-independent NAD glycohydrolase activity. Toxin-catalyzed NAD hydrolysis was increased synergistically by ATP and detergents or phospholipids; the zwitterionic detergent 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS) was more effective than the nonionic detergent Triton X-100 > lysophosphatidylcholine > phosphatidylcholine. The A0.5 for ATP in the presence of CHAPS was 2.6 μM; significantly higher concentrations of ATP were required for maximal activation in the presence of cholate or lysophosphatidylcholine. In CHAPS, NAD hydrolysis was enhanced by ATP > ADP > AMP > adenosine; ATP was more effective than MgATP or the nonhydrolyzable analogue adenyl-5'-yl imidodiphosphate. GTP and guanyl-5'-yl imidodiphosphate were less active than the corresponding adenine nucleotides. Activity in the presence of CHAPS and ATP was almost completely dependent on dithiothreitol; the A0.5 for dithiothreitol was significantly decreased by CHAPS alone and, to a greater extent, by CHAPS and ATP. To determine the site of action of ATP, CHAPS, and dithiothreitol, the enzymatic (S1) and binding components (B oligomer) were resolved by chromatography. The purified S1 subunit catalyzed the dithiothreitol-dependent hydrolysis of NAD; activity was enhanced by CHAPS but not ATP. The studies are consistent with the conclusion that adenine nucleotides, dithiothreitol, and CHAPS act on the toxin itself rather than on the substrate; adenine nucleotides appear to be involved in the activation of toxin but not the isolated catalytic unit.

Original languageEnglish (US)
Pages (from-to)2720-2725
Number of pages6
JournalBiochemistry
Volume25
Issue number9
DOIs
StatePublished - Feb 21 1986

ASJC Scopus subject areas

  • Biochemistry

Fingerprint Dive into the research topics of 'Stimulation of the Thiol-Dependent ADP-Ribosyltransferase and NAD Glycohydrolase Activities of Bordetella Pertussis Toxin by Adenine Nucleotides, Phospholipids, and Detergents'. Together they form a unique fingerprint.

Cite this