Characterization by electron paramagnetic resonance of the interactions of L-arginine and L-thiocitrulline with the heme cofactor region of nitric oxide synthase

J. C. Salerno, C. Frey, K. McMillan, R. F.B.S.S. Williams Masters, O. W. Griffith

Research output: Contribution to journalArticle

54 Scopus citations

Abstract

Nitric oxide synthase (NOS) catalyzes sequential NADPH- and O2-dependent mono-oxygenase reactions converting L-arginine to N(ω)-hydroxy-L-arginine and N(ω) hydroxy-L-arginine to citrulline and nitric oxide. The homodimeric enzyme contains one heme/monomer, and that cofactor is thought to mediate both partial reactions. Here we show by electron paramagnetic resonance spectroscopy that binding of substrate L-arginine to neuronal NOS perturbs the heme cofactor binding pocket without directly interacting as a sixth axial heine ligand; heme iron is exclusively high spin. In contrast, binding of L-thiocitrulline, a NOS inhibitor, produces both high and low spin iron spectra; L-thiocitrulline sulfur is a sixth axial heine ligand in one, but not all, of the low spin forms. The high spin forms of the L-thiocitrulline NOS complex display a distortion in the opposite direction to that caused by L-arginine binding. The findings elucidate the binding interactions of L- arginine and L-thiocitrulline to neuronal NOS and demonstrate that each causes a unique perturbation to the heine cofactor pocket of NOS.

Original languageEnglish (US)
Pages (from-to)27423-27428
Number of pages6
JournalJournal of Biological Chemistry
Volume270
Issue number46
DOIs
StatePublished - 1995

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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