The guanidino-methylated arginine analogue NG monomethyl-L-arginine (L-NMMA) has been the standard nitric oxide synthase inhibitor used to evaluate the role of the L-arginine:nitric oxide pathway. However, L-NMMA and other methylated arginine residues are also synthesised in vivo by the action of a family of enzymes known as protein arginine methyltransferases. Proteolysis of proteins containing methylated arginine residues releases free methylarginine residues into the cytosol from where they may pass out of the cell into plasma. Of the three known methylarginine residues produced in mammals only asymmetrically methylated forms (L-NMMA and asymmetric dimethylarginine (ADMA)) but not symmetrically methylated arginine (symmetric dimethylarginine (SDMA)) inhibit nitric oxide synthase (NOS). We and others have proposed that endogenously produced asymmetrically methylated arginines may modulate NO production and that the accumulation of these residues in disease states may contribute to pathology. The activity of the enzyme dimethylarginine dimethylaminohydrolase that metabolises asymmetric methylarginines may be of critical importance in affecting NO pathways in health or disease.

The enzyme dimethylarginine dimethylaminohydrolase (DDAH) hydrolyses asymmetrically methylated arginine residues that are endogenously produced inhibitors of nitric oxide synthases (NOS). We and others have proposed that DDAH activity is a key determinant of intracellular methylarginine concentrations and that factors that regulate the activity of DDAH may modulate nitric oxide (NO) production in vivo . We recently solved the crystal structure of a bacterial DDAH and identified a Cys-His-Glu catalytic triad [Murray-Rust, J., Leiper, J. M., McAlister, M., Phelan, J., Tilley, S., Santa Maria, J., Vallance, P. & McDonald, N. (2001) Nat. Struct. Biol. 8, 679–683]. The presence of a reactive cysteine residue (Cys-249) in the active site of DDAH raised the possibility that DDAH activity might be directly regulated by S -nitrosylation of this residue by NO. In the present study, we demonstrate that recombinant DDAH is reversibly inhibited after incubation with NO donors in vitro . Similarly mammalian DDAH in cytosolic extracts is also reversibly inhibited by NO donors. In cultured endothelial cells, heterologously expressed human DDAH II was S -nitrosylated after cytokine induced expression of the inducible NOS isoforms. The implication of these findings is that under certain conditions when NO generation increases, S -nitrosylation diminishes DDAH activity and this would be expected to lead to accumulation of asymmetric dimethylarginine and inhibition of NOS. This observation may help explain why expression of iNOS often leads to inhibition of activity of constitutively expressed NOS isozymes. We also identify Cys-His-Glu as a nitrosylation motif that is conserved in a family of arginine handling enzymes.

Background—: Proton pump inhibitors (PPIs) are gastric acid–suppressing agents widely prescribed for the treatment of gastroesophageal reflux disease. Recently, several studies in patients with acute coronary syndrome have raised the concern that use of PPIs in these patients may increase their risk of major adverse cardiovascular events. The mechanism of this possible adverse effect is not known. Whether the general population might also be at risk has not been addressed. Methods and Results—: Plasma asymmetrical dimethylarginine (ADMA) is an endogenous inhibitor of nitric oxide synthase. Elevated plasma ADMA is associated with increased risk for cardiovascular disease, likely because of its attenuation of the vasoprotective effects of endothelial nitric oxide synthase. We find that PPIs elevate plasma ADMA levels and reduce nitric oxide levels and endothelium-dependent vasodilation in a murine model and ex vivo human tissues. PPIs increase ADMA because they bind to and inhibit dimethylarginine dimethylaminohydrolase, the enzyme that degrades ADMA. Conclusions—: We present a plausible biological mechanism to explain the association of PPIs with increased major adverse cardiovascular events in patients with unstable coronary syndromes. Of concern, this adverse mechanism is also likely to extend to the general population using PPIs. This finding compels additional clinical investigations and pharmacovigilance directed toward understanding the cardiovascular risk associated with the use of the PPIs in the general population.