Metalloprotein-dependent decomposition of S-nitrosothiols: studies on the stabilization and measurement of S-nitrosothiols in tissues

Free Radical Biology & Medicine
Ali R ManiKevin Moore

Abstract

The stabilization of S-nitrosothiols is critical for the development of assays to measure their concentration in tissues. Low-molecular-weight S-nitrosothiols are unstable in tissue homogenates, even in the presence of thiol blockers or metal-ion chelators. The aim of this study was to try and stabilize low-molecular-weight S-nitrosothiols in tissue and gain insight into the mechanisms leading to their decomposition. Rat tissues (liver, kidney, heart, and brain) were perfused and homogenized in the presence of a thiol-blocking agent (N-ethylmaleimide) and a metal-ion chelator (DTPA). Incubation of liver homogenate with low-molecular-weight S-nitrosothiols (L-CysNO, D-CysNO, and GSNO) resulted in their rapid decomposition in a temperature-dependent manner as measured by chemiluminescence. The decomposition of L-CysNO requires a cytoplasmic factor, with activity greatest in liver > kidney > heart > brain > plasma, and is inhibitable by enzymatic proteolysis or heating to 80 degrees C, suggesting that a protein catalyzes the decomposition of S-nitrosothiols. The ability of liver homogenate to catalyze the decomposition of L-CysNO is up-regulated during endotoxemia and is dependent on oxygen, with the major product being nitrate. M...Continue Reading

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Citations

Apr 10, 2013·Journal of Physiology and Biochemistry·Asghar GhasemiSaleh Zahediasl
Jul 10, 2012·Antioxidants & Redox Signaling·Bradley A MaronJoseph Loscalzo
Jul 11, 2014·European Journal of Pharmacology·Golnar EftekhariAli R Mani
Oct 4, 2011·Clinical Biochemistry·Ignazio GrattaglianoGiuseppe Palasciano
Apr 3, 2009·Journal of Molecular and Cellular Cardiology·Sergiy M NadtochiyPaul S Brookes
Mar 18, 2010·British Journal of Pharmacology·M P Gordge, F Xiao
Jan 30, 2007·Hepatology : Official Journal of the American Association for the Study of Liver Diseases·Ignazio GrattaglianoGiuseppe Palasciano
Dec 21, 2006·The Biochemical Journal·Susannah E BellMichael P Gordge

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