BRAIN NITRIC-OXIDE SYNTHASE IS A BIOPTERIN-CONTAINING AND FLAVIN-CONTAINING MULTIFUNCTIONAL OXIDOREDUCTASE

Brain nitric oxide synthase is a Ca2+/calmodulin-regulated enzyme which converts L-arginine into NO. Enzymatic activity of this enzyme essentially depends on NADPH and is stimulated by tetrahydrobiopterin (H-4biopterin). We found that purified NO synthase contains enzyme-bound H-4biopterin, explaining the enzymatic activity observed in the absence of added cofactor. Together with the finding that H-4biopterin was effective at sub-stoichiometrical concentrations, these results indicate that NO synthase essentially depends on H-4biopterin as a cofactor which is recycled during enzymatic NO formation. We found that the purified enzyme also contains FAD, FMN and non-heme iron in equimolar amounts and exhibits striking activities, including a Ca2+/calmodulin-dependent NADPH oxidase activity, leading to the formation of hydrogen peroxide at suboptimal concentrations of L-arginine or H-4biopterin.