A point-mutated guanylyl cyclase with features of the YC-1-stimulated enzyme: implications for the YC-1 binding site?
Guanylyl cyclases (GCs) and adenylyl cyclases (ACs) play key roles in various signaling cascades and are structurally closely related. The crystal structure of a soluble AC revealed one binding site each for the substrate ATP and the activator forskolin. Recently, YC-1, a novel activator of the heterodimeric soluble GC (sGC), has been identified which acts like forskolin on AC. Here, we investigated the respective substrate and potential activator domains of sGC using point-mutated subunits. Whereas substitution of the conserved Cys-541 of the beta(1) subunit with serine led to an almost complete loss of activity, mutation of the respective homologue (Cys-596) in the alpha(1) subunit yielded an enzyme with an increased catalytic rate and higher sensitivity toward NO. This phenotype exhibits characteristics similar to those of the YC-1-treated wild-type enzyme. Conceivably, this domain which corresponds to the forskolin site of the ACs may comprise the binding site for YC-1.
Friebe
A
A
Institut für Pharmakologie, Freie Universität Berlin, Germany.
Russwurm
M
M
Mergia
E
E
Koesling
D
D
D000818:Animals; D001665:Binding Sites / Q000235:genetics; D020134:Catalytic Domain / Q000235:genetics; D002417:Cattle; D017124:Conserved Sequence / Q000235:genetics; D006152:Cyclic GMP / Q000378:metabolism; D019281:Dimerization; D020536:Enzyme Activators / Q000378:metabolism / Q000494:pharmacology*; D006160:Guanosine Triphosphate / Q000378:metabolism; D006162:Guanylate Cyclase / Q000235:genetics* / Q000378:metabolism*; D007191:Indazoles / Q000378:metabolism / Q000494:pharmacology*; D008274:Magnesium / Q000378:metabolism; D008345:Manganese / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009569:Nitric Oxide / Q000378:metabolism / Q000502:physiology; D017354:Point Mutation; D012995:Solubility
D020536:Enzyme Activators; D007191:Indazoles; C090937:3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole; D009569:Nitric Oxide; D008345:Manganese; D006160:Guanosine Triphosphate; D006162:Guanylate Cyclase; D006152:Cyclic GMP; D008274:Magnesium