1. The Km for NAD+ of cholera toxin working as an NAD+ glycohydrolase is 4 mM, and this is increased to about 50 mM in the presence of low-Mr ADP-ribose acceptors. Only molecules having both the adenine and nicotinamide moieties of NAD+ with minor alterations in the nicotinamide ring can be competitive inhibitors of this reaction. 2. This high Km for NAD+ is also reflected in the dissociation constant, Kd, which was determined by a variety of methods. 3. Results from equilibrium dialysis were subject to high error, but showed one binding site and a Kd of about 3 mM. 4. The A1 peptide of the toxin is digested by trypsin, and this digestion is completely prevented by concentrations of NAD+ above 50 mM. Measurement (by densitometric scanning of polyacrylamide-gel electrophoretograms) of the rate of tryptic digestion at different concentrations of NAD+ allowed a more accurate determination of Kd = 4.0 +/- 0.4 mM. Some analogues of NAD+ that are competitive inhibitors of the glycohydrolase reaction also prevented digestion.
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May 1987
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Research Article|
May 15 1987
Binding of NAD+ by cholera toxin
T S Galloway;
T S Galloway
1Department of Biochemistry, University of Edinburgh, U.K.
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S van Heyningen
S van Heyningen
1Department of Biochemistry, University of Edinburgh, U.K.
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Publisher: Portland Press Ltd
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© 1987 London: The Biochemical Society
1987
Biochem J (1987) 244 (1): 225–230.
Citation
T S Galloway, S van Heyningen; Binding of NAD+ by cholera toxin. Biochem J 15 May 1987; 244 (1): 225–230. doi: https://doi.org/10.1042/bj2440225
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