In vitro activation of soluble guanylyl cyclase and nitric oxide release: A comparison of NO donors and NO mimetics

Nitric oxide (NO) performs a central role in biological systems, binding to the heme site of soluble guanylyl cyclase (sGC), leading to enzyme activat ion and elevation of intracellular levels of cGMP. Organic nitrates, in par ticular, nitroglycerin (GTN), are clinically important nitrovasodilators th at function as NO-mimetics in biological systems. Comparison of sGC activat ion data with electrochemically measured rates of NO release for genuine NO donors, NONOates and nitrosothiols, yields an excellent correlation betwee n the EC50 for sGC activation and the rate constant for NO release, k(NO). However, activation of sGC by GTN and the nitrates has very different chara cteristics, including the requirement for specific added thiols, for exampl e, cysteine. The reaction of GTN with cysteine in anaerobic solution yields NO slowly, and NO release, measured by chemiluminescence detection, is que nched by added metal ion chelator. The generation of NO under aerobic condi tions is 100-fold slower than the anaerobic reaction. Furthermore. NO relea se from the reaction of GTN with cysteine in phosphate buffer is too slow t o account for sGC activation by GTN/cysteine. The slow rate of the chemical reaction to release NO suggests that nitrates can activate sGC by an NO-in dependent mechanism. In contrast to the genuine NO donors, GTN behaves as a partial agonist with respect to sGC activation, but in the presence of the allosteric sGC activator, YC-1, GTN exhibits full agonist activity.