title: Guanylyl cyclases A and B are asymmetric dimers that are allosterically activated by ATP binding to the catalytic domain.
abstract: It is not known how natriuretic peptides and adenosine triphosphate (ATP) activate guanylyl cyclase A (GC-A) and GC-B, which generate the second messenger cyclic guanosine monophosphate. We determined that natriuretic peptides increased the maximum rate of these enzymes >10-fold in a positive cooperative manner in the absence of ATP. In the absence of natriuretic peptides, ATP shifted substrate-velocity profiles from cooperative to linear but did not increase the affinity of GCs for the substrate guanosine triphosphate (GTP) since the Michaelis constant was unchanged. However, in the presence of natriuretic peptides, ATP competed with GTP for binding to an allosteric site, which enhanced the activation of GCs by decreasing the Michaelis constant. Thus, natriuretic peptide binding was required for communication of the allosteric activation signal to the catalytic site. The ability of ATP to activate GCs decreased and enzyme potency (a measure of sensitivity to stimulation) increased with increasing GTP concentrations. Point mutations in the purine-binding site of the catalytic domain abolished GC activity but not allosteric activation. Coexpression of inactive mutants produced half the activity expected for symmetric catalytic dimers. 2'-Deoxy-ATP and 2'-deoxy-GTP were poor allosteric activators, but 2'-deoxy-GTP was an effective substrate, consistent with distinct binding requirements for the allosteric and catalytic sites. We conclude that membrane GC domains are asymmetric homodimers with distinct and reciprocally regulated catalytic and allosteric sites that bind to GTP and ATP, respectively. These data define a new membrane GC activation model and provide evidence of a previously unidentified GC drug interaction site.
authors.affiliation: Department of Pharmacology, University of Minnesota, Minneapolis, MN 55455, USA.
authors.forename: Jerid W
authors.initials: JW
authors.lastname: Robinson
authors.forename: Lincoln R
authors.initials: LR
authors.lastname: Potter
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D000494:Allosteric Regulation / Q000502:physiology*; D000495:Allosteric Site / Q000235:genetics; D000818:Animals; D019281:Dimerization; D004789:Enzyme Activation / Q000502:physiology; D006160:Guanosine Triphosphate / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D008954:Models, Biological; D016296:Mutagenesis; D045265:Natriuretic Peptides / Q000378:metabolism*; D017354:Point Mutation / Q000235:genetics; D017461:Receptors, Atrial Natriuretic Factor / Q000737:chemistry / Q000378:metabolism*; D012044:Regression Analysis; D015290:Second Messenger Systems / Q000502:physiology*; D000099242:Atrial Natriuretic Factor Receptor B
chemical_list: D000255:Adenosine Triphosphate; D006160:Guanosine Triphosphate; D045265:Natriuretic Peptides; D017461:Receptors, Atrial Natriuretic Factor; D000099242:Atrial Natriuretic Factor Receptor B; C091715:atrial natriuretic factor receptor A