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Nitric oxide and NAD-dependent protein modification
1Laboratory of Cellular Metabolism, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892.
Molecular and Cellular Biochemistry
|September 1, 1994
Summary
Nitric oxide (NO) does not stimulate ADP-ribosylation. Instead, NO modifies glyceraldehyde-3-phosphate dehydrogenase (GAPDH) by covalently binding the entire NAD molecule, possibly due to S-nitrosylation.
Area of Science:
- Biochemistry
- Molecular Biology
- Enzymology
Background:
- Nitric oxide (NO) was previously hypothesized to regulate intracellular ADP-ribosylation.
- This hypothesis was based on protein radiolabelling studies using [32P]NAD and NO in tissue homogenates.
Purpose of the Study:
- To investigate the mechanism of NO-mediated protein modification.
- To determine if NO stimulates an endogenous ADP-ribosyltransferase, specifically targeting glyceraldehyde-3-phosphate dehydrogenase (GAPDH).
Main Methods:
- Replication of NO-stimulated modification in a defined system using purified GAPDH.
- Biochemical characterization of the NO-stimulated, NAD-dependent modification of GAPDH.
Main Results:
- The NO-stimulated modification of GAPDH was confirmed to be the covalent binding of the entire NAD molecule to the enzyme, not ADP-ribosylation.
- GAPDH is known to be stoichiometrically S-nitrosylated.
Conclusions:
- The initial hypothesis of NO-stimulation of ADP-ribosylation is disproven.
- NO's role in modifying GAPDH involves the covalent attachment of NAD, potentially by inducing S-nitrosylation of a cysteine residue in the enzyme's active site, thereby altering its chemical reactivity.