Tetrahydrobiopterin-deficient nitric oxide synthase has a modified heme environment and forms a cytochrome P-420 analogue

Jianling Wang, Dennis J. Stuehr, Denis L. Rousseau

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Abstract

Optical absorption and resonance Raman spectra of neuronal nitric oxide synthase (b-NOS) isolated in the absence of tetrahydrobiopterin demonstrate that the enzyme preparation is very unstable. This unstable form of the enzyme has properties analogous to those of cytochrome P-420cam, an inactive form of cytochrome P-450cam. Although cysteine is preserved as the proximal ligand in both the ferric and ferrous forms of unstable b-NOS, the lack of tetrahydrobiopterin significantly increases the hexacoordinate low-spin fraction of the heme content, resulting in a loss of the enzymatic activity. Upon the addition of CO, the unstable b-NOS converts from a species exhibiting a Soret absorption maximum at 443 nm, as reported for the CO adducts of stable b-NOS and cytochrome P-450cam, to a species with a Soret maximum at 421 nm. The resonance Raman spectrum of the 421-nm form is the same as those of CO-bound myoglobin at low pH and CO-bound cytochrome P-420cam. The heme in this form of the enzyme is coordinated by a weaker ligand than thiolate; histidine coordination in the CO-bound form of the P-420-like species of NOS is consistent with all of the available data. A similar unstable form of the macrophage (i-NOS) enzyme was also detected. Not only does the heme pocket of NOS have the same coordination as cytochrome P-450 in its stable form, but the partially denatured form has the same properties as cytochrome P-420, the inactive form of cytochrome P-450. Among other possible roles, tetrahydrobiopterin may play a significant role in the stabilization of the active enzyme.

Original languageEnglish (US)
Pages (from-to)7080-7087
Number of pages8
JournalBiochemistry
Volume34
Issue number21
StatePublished - 1995
Externally publishedYes

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Carbon Monoxide
Heme
Nitric Oxide Synthase
Cytochromes
Enzymes
Cytochrome P-450 Enzyme System
Raman scattering
Ligands
Nitric Oxide Synthase Type I
Myoglobin
Macrophages
Histidine
Light absorption
Cysteine
Stabilization
sapropterin
cytochrome P420

ASJC Scopus subject areas

  • Biochemistry

Cite this

Tetrahydrobiopterin-deficient nitric oxide synthase has a modified heme environment and forms a cytochrome P-420 analogue. / Wang, Jianling; Stuehr, Dennis J.; Rousseau, Denis L.

In: Biochemistry, Vol. 34, No. 21, 1995, p. 7080-7087.

Research output: Contribution to journalArticle

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