title: Mutations in Tyr808 reveal a potential auto-inhibitory mechanism of guanylate cyclase-B regulation.
abstract: In this study, Tyr808 in GC-B (guanylate cyclase-B), a receptor of the CNP (C-type natriuretic peptide), has been shown to be a critical regulator of GC-B activity. In searching for phosphorylation sites that could account for suppression of GC-B activity by S1P (sphingosine-1-phosphate), mutations were introduced into several candidate serine/threonine and tyrosine residues. Although no novel phosphorylation sites that influenced the suppression of GC-B were identified, experiments revealed that mutations in Tyr808 markedly enhanced GC-B activity. CNP-stimulated activities of the Y808F and Y808A mutants were greater than 30-fold and 70-fold higher, respectively, than that of WT (wild-type) GC-B. The Y808E and Y808S mutants were constitutively active, expressing 270-fold higher activity without CNP stimulation than WT GC-B. Those mutations also influenced the sensitivity of GC-B to a variety of inhibitors, including S1P, Na3VO4 and PMA. Y808A, Y808E and Y808S mutations markedly weakened S1P- and Na3VO4-dependent suppression of GC-B activity, whereas Y808E and Y808S mutations rather elevated cGMP production. Tyr808 is conserved in all membrane-bound GCs and located in the niche domain showing sequence similarity to a partial fragment of the HNOBA (haem nitric oxide binding associated) domain, which is found in soluble GC and in bacterial haem-binding kinases. This finding provides new insight into the activation mechanism of GCs.
authors.affiliation: Department of Pharmacology, University of Texas Southwestern Medical Center at Dallas, TX, USA. Takeshi.Katafuchi@UTSouthwestern.edu
authors.forename: Takeshi
authors.initials: T
authors.lastname: Katafuchi
mesh_terms: D004789:Enzyme Activation; D006367:HeLa Cells; D006801:Humans; D010766:Phosphorylation; D017354:Point Mutation; D017461:Receptors, Atrial Natriuretic Factor / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism*; D014443:Tyrosine / Q000235:genetics* / Q000378:metabolism; D000099242:Atrial Natriuretic Factor Receptor B
chemical_list: D017461:Receptors, Atrial Natriuretic Factor; D014443:Tyrosine; D000099242:Atrial Natriuretic Factor Receptor B