NO activation of guanylyl cyclase

Abstract

Nitric oxide (NO)-sensitive guanylyl-cyclase (GC) is the most important receptor for the signaling molecule NO. Activation of the enzyme is brought about by binding of NO to the prosthetic heme group. By monitoring NObinding and catalytic activity simultaneously, we show that NO activates GC only if the reaction products of the enzyme are present. NO-binding in the absence of the products did not activate the enzyme, but yielded a nonactivated species with the spectral characteristics of the active form. Conversion of the nonactivated into the activated conformation of the enzyme required the simultaneous presence of NO and the reaction products. Furthermore, the products magnesium/cGMP/pyrophosphate promoted the release of the histidine-iron bond during NO-binding, indicating reciprocal communication of the catalytic and ligand-binding domains. Based on these observations, we present a model that proposes two NO-bound states of the enzyme: an active state formed in the presence of the products and a nonactivated state. The model not only covers the data reported here but also consolidates results from previous studies on NO-binding and dissociation/deactivation of GC.

Tonic and acute nitric oxide signaling through soluble guanylate cyclase is mediated by nonheme nitric oxide, ATP, and GTP

“…This finding indicates that conversion of the six-coordinate intermediate to the five-coordinate complex is accelerated by GTP. Similar data have been reported (21). However, when ATP is included with GTP, a peak at 420 nm appears.…”

“…This finding is consistent with a recent report in which a low-activity species of sGC with NO bound to the heme was described. In this report, when excess NO was removed from sGC, the activity of the resulting ferrous-nitrosyl enzyme was 10-20% of full activity (21). Together, these observations indicate that the formation of ferrous-nitrosyl sGC is not always sufficient for full enzyme activation.…”

“…The sGC heme-NO complex is extremely stable, inconsistent with a rapid deactivation profile in vivo. Recent data also suggest that full activation of sGC by NO at the heme requires high levels of the cyclase reaction products [cGMP and͞or pyrophosphate (PP i )] before NO binding (21). Furthermore, ATP at physiological concentrations inhibits sGC activity by as much as 50% (22,23).…”

Structural insights into the molecular mechanism of H‐NOX activation

“…[1][2][3] In sGC, NO-mediated breakage of the Fe-His bond leads to increased catalytic activity. Recently, heme domains related to sGC were identified in prokaryotes and were termed Heme-Nitric oxide/OXygen binding (H-NOX) domains.…”

“…The second class consists of H-NOX domains fused to methyl-accepting chemotaxis proteins. 6,4,7 Homology to the sGC heme domain as well as genomic location adjacent to putative signaling proteins suggests that H-NOX domains likely serve as O 2 and NO sensors in prokaryotes. 4,7 Consistent with this hypothesis, the ferrous NO complex of the H-NOX from the facultative aerobe, Shewanella oneidensis (So H-NOX), which belongs to the first class of H-NOX proteins, inhibits autophosphorylation of a histidine kinase encoded in the same operon, whereas the ferrous unliganded protein has no effect on kinase activity.…”