Genetically tailored atrial natriuretic factor-dependent guanylate cyclase Immunological and functional identity with 180 kDa membrane guanylate cyclase and ATP signaling site

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Abstract
Biochemical and immunological studies have established that one of the signal transducers of atrial natriuretic factor (ANF) is a 180 kDa membrane guanylate cyclase (180 kDa mGC), which is also an ANF receptor; obligatory in the transduction process is an intervening ATP-regulated step, but its mechanism is not known. GCα is a newly discovered member of the guanylate cyclase family whose activity is independent of the known natriuretic peptides, and the enzyme is not an ANF receptor. The genetically tailored GCα, GCα-DmutGln338 Leu364, however, is not only a guanylate cyclase but also an ANF receptor and is structurally and functionally identical to the cloned wild-type ANF receptor guanylate cyclase, GC-A. We now report that the ANF-dependent guanylate cyclase activity in the particulate fractions of cells transfected with GCα-DmutGln338 Leu364 was inhibited by the 180 kDa mGC polyclonal antibody, and with this antibody probe it was possible to purify the 130 kDa expressed receptor; the hormone-dependent cyclase activity of this receptor was exclusively dependent upon ATP; and through site-directed mutational studies with GCα mutants, the signaling sequence that defines ATP binding site was identified. We thus conclude that 180 kDa mGC and the mutant protein are immunologically similar; both proteins are linked to the ANF signal in the generation of cyclic GMP synthesis; and in both the ligand binding and catalytic activities are bridged through a defined ATP binding module