Regulation of human neutrophil guanylate cyclase by metal ions, free radicals and the muscarinic cholinergic receptor

Abstract

We have examined the properties of soluble guanylate cyclase activity in the human neutrophil. The enzyme showed complex regulation by metal ions. A 10-fold higher activity was observed in the presence of Mn^(2+) than Mg^(2+), while Ca^(2+) caused an increase in activity only in the presence of Mg^(2+) ion. Sodium nitroprusside (SNP), azide and hydrogen peroxide were activators of the enzyme. Dithiothreitol blocked the activation by SNP, suggesting the involvement of thiol groups in the activation process. Carbachol acting through the muscarinic cholinergic receptor caused a dose-dependent activation, which was blocked by atropine. Higher concns of carbachol were required to activate guanylate cyclase than were required for the modulation of enzyme release elicited by N-fortnyl-L-methionyl-L-leucyl-L-phenylalanine. Nordihydroguaracetic acid inhibited carbachol stimulation of guanylate cyclase. By contrast, trifluoperazine (TFP), a calmodulin antagonist, caused a biphasic modulation of basal activity in the presence or absence of carbachol. Our results indicate that: (a) allosteric interactions of metal ions are important to the regulation of the enzyme, (b) the free radical nitroxide as well as hydrogen peroxide enhances enzyme activity, (c) agonist occupancy of the muscarinic cholinergic receptor activates neutrophil guanylate cyclase probably through a mechanism involving calcium influx and the activation of the lipoxygenase pathway, and (d) a TFP-sensitive site (possibly calmodulin) is involved in the selective regulation of basal enzyme activity.

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