Low-temperature optical absorption spectra suggest a redox role for tetrahydrobiopterin in both steps of nitric oxide synthase catalysis.

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Gorren AC, Bec N, Schrammel A, Werner ER, Lange R, Mayer B

Low-temperature optical absorption spectra suggest a redox role for tetrahydrobiopterin in both steps of nitric oxide synthase catalysis.

Biochemistry. 2000 Sep 26;39(38):11763-70.

PubMed ID

10995244 [ View in PubMed

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Abstract

To investigate the role of tetrahydrobiopterin (BH4) in the catalytic mechanism of nitric oxide synthase (NOS), we analyzed the spectral changes following addition of oxygen to the reduced oxygenase domain of endothelial nitric oxide synthase (NOS) in the presence of different pteridines at -30 degrees C. In the presence of N(G)-hydroxy-L-arginine (NOHLA) and BH4 or 5-methyl-BH4, both of which support NO synthesis, the first observable species were mixtures of high-spin ferric NOS (395 nm), ferric NO-heme (439 nm), and the oxyferrous complex (417 nm). With Arg, no clear intermediates could be observed under the same conditions. In the presence of the BH4-competitive inhibitor 7,8-dihydrobiopterin (BH2), intermediates with maxima at 417 and 425 nm were formed in the presence of Arg and NOHLA, respectively. In the presence of 4-amino-BH4, the maxima of the intermediates with Arg and NOHLA were at 431 and 423 nm, respectively. We ascribe all four spectra to oxyferrous heme complexes. The intermediates observed in this study slowly decayed to the high-spin ferric state at -30 degrees C, except for those formed in the presence of 4-amino-BH4, which required warming to room temperature for regeneration of high-spin ferric NOS; with Arg, regeneration remained incomplete. From these observations, we draw several conclusions. (1) BH4 is required for reductive oxygen activation, probably as a transient one-electron donor, not only in the reaction with Arg but also with NOHLA; (2) in the absence of redox-active pterins, reductive oxygen activation does not occur, which results in accumulation of the oxyferrous complex; (3) the spectral properties of the oxyferrous complex are affected by the presence and identity of the substrate; (4) the slow and incomplete formation of high-spin ferric heme with 4-amino-BH4 suggests a structural cause for inhibition of NOS activity by this pteridine.

DrugBank Data that Cites this Article

Drug Targets

Drug	Target	Kind	Organism	Pharmacological Action	Actions
Sapropterin	Nitric oxide synthase, endothelial	Protein	Humans	Yes	Cofactor	Details