Inhibition of GAPDH by S-nitrosothiols and nitric oxide in cellular environment and in isolation
Broniowska, KA; Hogg, N
HERO ID
975895
Reference Type
Journal Article
Subtype
Abstract
Year
2008
Language
English
| HERO ID | 975895 |
|---|---|
| Material Type | Abstract |
| In Press | No |
| Year | 2008 |
| Title | Inhibition of GAPDH by S-nitrosothiols and nitric oxide in cellular environment and in isolation |
| Authors | Broniowska, KA; Hogg, N |
| Journal | Free Radical Biology and Medicine |
| Volume | 45 |
| Issue | Suppl. |
| Page Numbers | S110-S110 |
| Abstract | S-Nitrosothiols are molecules involved in nitric oxide (NO) biology and often thought to be NO reservoirs. They represent a noncGMP-dependent route of transducing NO activity via modification of critical thiols. Although S-nitrosation of key cysteine residues has been suggested to represent a novel redox-based signaling mechanism, the exact mechanisms of S-nitrosothiols formation under (patho)physiological conditions and the signaling specificity have yet to be established. Here, we examined the sensitivity of GAPDH to inhibition by CysNO and NO inside the cell and in isolation. Bovine aortic endothelial cells and purified GAPDH preparations were treated with S-nitrosocysteine (CysNO) or NONOate-type NO donors, and enzymatic activity was monitored by following NADH oxidation. Intracellular GAPDH was irreversibly inhibited upon CysNO administration, whereas treatment with NO resulted in a DTT-reversible inhibition of the enzyme. Purified GAPDH was inhibited by both CysNO and NO. Interestingly, the DTT-reversibility of inhibition was diametrically opposite to that observed in cells. CysNO-dependent inhibition was reversed with DTT. This did not occur with NO-dependent inhibition. However, NO inhibited purified GAPDH in a DTT-reversible way in the presence of GSH. in addition, enzyme that had been pre-treated with CysNO and then treated with NO, also recovered activity upon incubation with DTT. Our data suggest that in response to CysNO treatment, cellular GAPDH undergoes S-nitrosation that is followed by further reactions resulting in an irreversible inhibition, whereas NO inhibits the enzyme via an oxidative mechanism that does not involve S-nitrosation. In summary, our data show that GAPDH is a sensitive target for CysNO- and NO-dependent inhibition, however these two agents inhibit the enzyme via different mechanisms both inside the cell and in isolation. Additionally, discrepancies observed between the cellular system and purified protein strongly imply that the intracellular milieu is actively involved in dictating the mechanism of inhibition. |
| Wosid | WOS:000260867900311 |
| Url | http://www.sciencedirect.com/science/article/pii/S089158490800628X |
| Is Certified Translation | No |
| Dupe Override | No |
| Conference Location | Indianapolis, IN |
| Conference Name | Society for Free Radical Biology and Medicine 15th Annual Meeting |
| Conference Date | November 19-23, 2008 |
| Is Public | Yes |
| Language Text | English |
| Relationship(s) |
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