Inactivation of nitric oxide synthase isoforms by diaminoguanidine and NG-amino-L-arginine.
Diaminoguanidine (DAG) and NG-amino-L-arginine each produced a time- and concentration-dependent inactivation of the citrulline-forming activity of all three NOS isoforms. DAG inactivates both the NADPH-oxidase and the citrulline-forming activities of GH3 pituitary nNOS while NG-amino-L-arginine inactivates only its citrulline-forming activity. The inactivation by DAG of GH3 nNOS NADPH-oxidase and citrulline forming activities is stimulated by (6R)-5,6,7,8-tetrahydrobiopterin (BH4) cofactor, follows pseudo-first-order kinetics and is not substrate saturable. DAG-induced inactivation of the citrulline-forming activity for the iNOS and eNOS isoforms displayed maximal inactivation rates of 0.37 and 0.14 min-1 and Ki values of 385 and 670 microM, respectively. At 1 mM DAG and saturating BH4, half-times of inactivation of 0.7, 8, and 2 min were observed for the nNOS, eNOS, and iNOS isoforms, respectively. NG-Amino-L-arginine-induced inactivation of the citrulline-forming activity of the nNOS, iNOS, and eNOS isoforms displayed maximal inactivation rates of 0.35, 0.26, and 0.53 min-1 and Ki values of 0.3, 3, and 2.5 microM, respectively. The inactivation of the NOS activities by both DAG and NG-amino-L-arginine in preincubations required the presence of oxygen and Ca2+, consistent with an inactivation mechanism that requires active metabolism by NOS. Methylguanidine and 1,1-dimethylguanidine exhibited a reversible inhibition pattern in contrast to all three NOS isoforms. Neither agent exhibited significant isoform selectivity.
Wolff
D J
DJ
Department of Pharmacology, University of Medicine and Dentistry of New Jersey, Robert Wood Johnson Medical School, Piscataway 08854, USA.
Lubeskie
A
A
D000818:Animals; D001120:Arginine / Q000031:analogs & derivatives* / Q000494:pharmacology; D002417:Cattle; D002956:Citrulline / Q000096:biosynthesis; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006146:Guanidines / Q000494:pharmacology*; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000037:antagonists & inhibitors*; D008760:Methylguanidine / Q000031:analogs & derivatives / Q000494:pharmacology; D051379:Mice; D009247:NADH, NADPH Oxidoreductases / Q000037:antagonists & inhibitors; D019255:NADPH Oxidases; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors*; D010084:Oxidation-Reduction; D051381:Rats
D004791:Enzyme Inhibitors; D006146:Guanidines; D007527:Isoenzymes; D002956:Citrulline; C070130:diaminoguanidine; C063904:N(G)-aminoarginine; D008760:Methylguanidine; C042632:N,N-dimethylguanidine; D001120:Arginine; D019001:Nitric Oxide Synthase; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases