X-ray absorption spectroscopic analysis of the high-spin ferriheme site in substrate-bound neuronal nitric-oxide synthase

J Biochem. 2001 Aug;130(2):191-8. doi: 10.1093/oxfordjournals.jbchem.a002972.


Nitric oxide synthase (NOS) catalyzes the conversion of L-arginine to citrulline and nitric oxide through two stepwise oxygenation reactions involving N(omega)-hydroxy-L-arginine, an enzyme-bound intermediate. The N(omega)-hydroxy-L-arginine- and arginine-bound NOS ferriheme centers show distinct, high-spin electron paramagnetic resonance signals. Iron X-ray absorption spectroscopy (XAS) has been used to examine the structure of the ferriheme site in the N(omega)-hydroxy-L-arginine-bound full-length neuronal NOS in the presence of (6R)-5,6,7,8-tetrahydro-L-biopterin. Iron XAS shows that the high-spin ferriheme sites in the N(omega)-hydroxy-L-arginine- and arginine-bound forms are strikingly similar, both being coordinated by the heme and an axial thiolate ligand, with an Fe-S distance of ca. 2.29 A. Cu(2+) inhibition slightly affects the spin-state equilibrium, but causes no XAS-detectable changes in the immediate ferriheme coordination environment of neuronal NOS. The structure and ligand geometry of the high-spin ferriheme in arginine-bound neuronal NOS are essentially identical to those of the N(omega)-hydroxy-L-arginine-bound form and only slightly affected by the divalent cation inhibitor of constitutive NOS.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Arginine / analogs & derivatives*
  • Arginine / chemistry
  • Arginine / metabolism
  • Biopterin / analogs & derivatives*
  • Biopterin / metabolism
  • Catalytic Domain
  • Cations, Divalent / pharmacology
  • Copper / metabolism
  • Electron Spin Resonance Spectroscopy
  • Enzyme Inhibitors / pharmacology
  • Heme / chemistry
  • Heme / metabolism
  • Humans
  • Ligands
  • Mice
  • Nitric Oxide / metabolism
  • Nitric Oxide Synthase / antagonists & inhibitors
  • Nitric Oxide Synthase / chemistry*
  • Nitric Oxide Synthase / metabolism*
  • Nitric Oxide Synthase Type I
  • Spectrum Analysis / methods*
  • X-Rays


  • Cations, Divalent
  • Enzyme Inhibitors
  • Ligands
  • Biopterin
  • Nitric Oxide
  • Heme
  • N(omega)-hydroxyarginine
  • Copper
  • Arginine
  • NOS1 protein, human
  • Nitric Oxide Synthase
  • Nitric Oxide Synthase Type I
  • Nos1 protein, mouse
  • sapropterin