Understanding the Structural Aspects of Neuronal Nitric Oxide Synthase (NOS) Using Microdissection by Molecular Cloning Techniques

Molecular Dissection of Neuronal NOS
  • B. S. S. Masters
  • K. McMillan
  • J. Nishimura
  • P. Martasek
  • L. J. Roman
  • E. Sheta
  • S. S. Gross
  • J. Salerno
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 387)


The neuronal isoform (Type I) of nitric oxide synthase (NOS) requires Ca+2/calmodulin to catalyze the formation of NO● and citrulline from L-arginine (1) and molecular oxygen (2) with reducing equivalents from NADPH. The overall reaction is a five-electron process involving two successive monooxygenation steps with the obligatory formation of N-hydroxy-L-arginine as the oxygenated intermediate. The neuronal NOS shares with the other two known isoforms the unusual property, for a mammalian enzyme, of containing iron protoporphyrin IX (3–6), FAD and FMN (3, 7–9), and tetrahydrobiopterin (9). Bredt, et al. (10) had previously demonstrated the remarkable (58%) sequence similarity of rat brain NOS to rat liver NADPH-cytochrome P450 reductase and determined that the C-terminal 641 amino acids of NOS display consensus regions for both flavin and nucleotide binding. It was reported that carbon monoxide (CO) inhibited both macrophage and neuronal NOS activity (3–6) and these various preparations exhibited reduced-CO difference spectra with absorbance maxima in the region of 445 nm, a property shared with the members of the cytochrome P450 family. However, the low degree of sequence similarity to any of the more than 200 members of this family and other CO-binding, oxygenating heme proteins, such as chloroperoxidase or Bacillus megaterium P450 (BM-3), suggests that the homology may end with the sharing of a cysteine thiolate ligand to the heme iron at the fifth axial position. Recent studies have established that this involves cysteine415 of the neuronal NOS (11, 12), as suggested originally by McMillan, et al. (3), and cysteine184 of the endothelial NOS (13) as determined by site-directed mutagenesis of the holoenzyme (11,13) and the heme domain (12), respectively.


Bacillus Megaterium Heme Protein Calmodulin Binding Cytochrome P450 Family Fatty Acid Hydroxylation 
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Copyright information

© Springer Science+Business Media New York 1996

Authors and Affiliations

  • B. S. S. Masters
    • 1
  • K. McMillan
    • 1
  • J. Nishimura
    • 1
  • P. Martasek
    • 1
  • L. J. Roman
    • 1
  • E. Sheta
    • 2
  • S. S. Gross
    • 3
  • J. Salerno
    • 4
  1. 1.Department of BiochemistryThe University of Texas Health Science Center at San AntonioSan AntonioUSA
  2. 2.Department of Biochemistry Faculty of ScienceAlexandria UniversityAlexandriaEgypt
  3. 3.Department of PharmacologyCornell University Medical CenterNew YorkUSA
  4. 4.Department of BiologyRensselaer Polytechnic InstituteTroyUSA

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