Tonic and acute nitric oxide signaling through soluble guanylate cyclase: Roles of non-heme nitric oxide, ATP, and globins.

Nitric oxide (NO) mediates blood vessel relaxation, complex aspects of myocardial function, perfusion and function of all major organs, synaptic plasticity in the brain, platelet aggregation, skin function, and numerous other physiological processes, by targeting and activating soluble guanylate cyclase (sGC). In the existing activation model, NO binds to the heme of sGC, forming a 6-coordinate intermediate. The rate of conversion of this intermediate to the final 5-coordinate ferrous-nitrosyl species depends on the concentration of NO; thus, non-heme NO somehow accelerates rupture of the proximal histidine-iron bond. Breaking of this bond is thought to result in a conformational change in the catalytic domain of the enzyme, accelerating the basal rate of conversion of GTP to 3',5'-cyclic GMP several hundred fold. cGMP triggers downstream cascades that result in the physiological responses to NO. Dysregulation of NO signaling, then, contributes to many types of disease state, from erectile dysfunction and heart disease, to neurodegeneration, stroke, hypertension, and gastrointestinal disease, to name a few. In vivo and ex vivo tissue studies of NO have revealed two fundamentally distinct and paradoxical signaling modes: tonic and acute. Tonic NO describes the continual low-level production of NO that elicits a long-lasting low-level cGMP signal. An acute burst of NO synthesis causes a rapid and transient rise in cGMP levels, resulting in processes such as vasodilation. These data, which describe two separate effects of NO, cannot be explained by the established binary model for NO regulation of sGC: that NO activates sGC solely by binding to its heme co-factor. To explore this discrepancy, I studied the kinetics of NO-heme dissociation and enzyme deactivation. In the presence of physiological concentrations of ATP and GTP, NO dissociation from the sGC heme is ∼500 times slower than the rate of enzyme deactivation in vitro. Deactivated sGC still has NO bound to the heme, and full activation requires additional NO. Intracellular globins trap NO in vivo, diminishing the acute response to NO; they also form a complex with sGC and accelerate dissociation of NO from the heme. We propose an activation model where, in the presence of both ATP and GTP, tonic NO forms a stable heme complex with low sGC activity; acute production of NO transiently and fully activates this NO-bound sGC. Globins accelerate deactivation of both species of NO-activated sGC.Ph.D.BiochemistryPure SciencesUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/125317/2/3192596.pd