title: Nitric oxide synthase is a cytochrome P-450 type hemoprotein.
abstract: Nitric oxide has emerged as an important mammalian metabolic intermediate involved in critical physiological functions such as vasodilation, neuronal transmission, and cytostasis. Nitric oxide synthase (NOS) catalyzes the five-electron oxidation of L-arginine to citrulline and nitric oxide. Cosubstrates for the reaction include molecular oxygen and NADPH. In addition, there is a requirement for tetrahydrobiopterin. NOS also contains the coenzymes FAD and FMN and demonstrates significant amino acid sequence homology to NADPH-cytochrome P-450 reductase. Herein we report the identification of the inducible macrophage NOS as a cytochrome P-450 type hemoprotein. The pyridine hemochrome assay showed that the NOS contained a bound protoporphyrin IX heme. The reduced carbon monoxide binding spectrum shows an absorption maximum at 447 nm indicative of a cytochrome P-450 hemoprotein. A mixture of carbon monoxide and oxygen (80%/20%) potently inhibited the reaction (73-79%), showing that the heme functions directly in the oxidative conversion of L-arginine to nitric oxide and citrulline. Additionally, partially purified NOS from rat cerebellum was inhibited by CO, suggesting that this isoform may also contain a P-450-type heme. NOS is the first example of a soluble cytochrome P-450 in eukaryotes. In addition, the presence of FAD and FMN indicates that this is the first catalytically self-sufficient mammalian P-450 enzyme, containing both a reductase and a heme domain on the same polypeptide.
authors.affiliation: Department of Biological Chemistry, Medical School, University of Michigan, Ann Arbor 48109-1065.
authors.forename: K A
authors.initials: KA
authors.lastname: White
authors.forename: M A
authors.initials: MA
authors.lastname: Marletta
mesh_terms: D000594:Amino Acid Oxidoreductases / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D001120:Arginine / Q000378:metabolism; D002248:Carbon Monoxide / Q000494:pharmacology; D002531:Cerebellum / Q000201:enzymology; D002956:Citrulline / Q000378:metabolism; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D006420:Hemeproteins / Q000302:isolation & purification / Q000378:metabolism*; D006900:Hydroxylation; D007700:Kinetics; D008264:Macrophages / Q000201:enzymology; D051379:Mice; D019001:Nitric Oxide Synthase; D013053:Spectrophotometry
chemical_list: D006420:Hemeproteins; D002956:Citrulline; D002248:Carbon Monoxide; D003577:Cytochrome P-450 Enzyme System; D001120:Arginine; D019001:Nitric Oxide Synthase; D000594:Amino Acid Oxidoreductases