Nucleotidyl cyclase activity of particulate guanylyl cyclase A: comparison with particulate guanylyl cyclases E and F, soluble guanylyl cyclase and bacterial adenylyl cyclases CyaA and edema factor.
Guanylyl cyclases (GCs) regulate many physiological processes by catalyzing the synthesis of the second messenger cGMP. The GC family consists of seven particulate GCs (pGCs) and a nitric oxide-activated soluble GC (sGC). Rat sGC α1β1 possesses much broader substrate specificity than previously assumed. Moreover, the exotoxins CyaA from Bordetella pertussis and edema factor (EF) from Bacillus anthracis possess nucleotidyl cyclase (NC) activity. pGC-A is a natriuretic peptide-activated homodimer with two catalytic sites that act cooperatively. Here, we studied the NC activity of rat pGC-A in membranes of stably transfected HEK293 cells using a highly sensitive and specific HPLC-MS/MS technique. GTP and ITP were effective, and ATP and XTP were only poor, pGC-A substrates. In contrast to sGC, pGC-A did not use CTP and UTP as substrates. pGC-E and pGC-F expressed in bovine rod outer segment membranes used only GTP as substrate. In intact HEK293 cells, pGC-A generated only cGMP. In contrast to pGCs, EF and CyaA showed very broad substrate-specificity. In conclusion, NCs exhibit different substrate-specificities, arguing against substrate-leakiness of enzymes and pointing to distinct physiological functions of cyclic purine and pyrimidine nucleotides.
Institute of Pharmacology, Hannover Medical School, Hannover, Germany.
Kerstin Y
KY
Beste
Corinna M
CM
Spangler
Heike
H
Burhenne
Karl-Wilhelm
KW
Koch
Yuequan
Y
Shen
Wei-Jen
WJ
Tang
Volkhard
V
Kaever
Roland
R
Seifert
D037361:Adenylate Cyclase Toxin / Q000378:metabolism; D000818:Animals; D000942:Antigens, Bacterial / Q000378:metabolism; D001427:Bacterial Toxins / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D006152:Cyclic GMP; D004789:Enzyme Activation; D006162:Guanylate Cyclase / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D007700:Kinetics; D051381:Rats; D017461:Receptors, Atrial Natriuretic Factor / Q000235:genetics / Q000378:metabolism*; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism; D000071756:Soluble Guanylyl Cyclase; D013379:Substrate Specificity
D037361:Adenylate Cyclase Toxin; D000942:Antigens, Bacterial; D001427:Bacterial Toxins; D018160:Receptors, Cytoplasmic and Nuclear; C030325:anthrax toxin; D006162:Guanylate Cyclase; D017461:Receptors, Atrial Natriuretic Factor; D000071756:Soluble Guanylyl Cyclase; C091715:atrial natriuretic factor receptor A; D006152:Cyclic GMP