Resonance Raman detection of the Fe-S bond in endothelial nitric oxide synthase

Johannes Schelvis, Vladimir Berka, Gerald T. Babcock, Ah Lim Tsai

Research output: Contribution to journalArticle

35 Citations (Scopus)

Abstract

We report the first low-frequency resonance Raman spectra of ferric endothelial nitric oxide synthase (eNOS) holoenzyme, including the frequency of the Fe-S vibration in the presence of the substrate L-arginine. This is the first direct measurement of the strength of the Fe-S bond in NOS. The Fe-S vibration is observed at 338 cm-1 with excitation at 363.8 nm. The assignment of this band to the Fe-S stretching vibration was confirmed by the observation of isotopic shifts in eNOS reconstituted with 54Fe- and 57Fe-labeled hemin. Furthermore, the frequency of this vibration is close to those observed in cytochrome P450cam and chloroperoxidase (CPO). The frequency of this vibration is lower in eNOS than in P450cam and CPO, which can be explained by differences in hydrogen bonding to the proximal cysteine heme ligand. On addition of substrate to eNOS, we also observe several low-frequency vibrations, which are associated with the heme pyrrole groups. The enhancement of these vibrations suggests that substrate binding results in protein-mediated changes of the heme geometry, which may provide the protein with an additional tuning element for the redox potential of the heme iron. The implications of our findings for the function of eNOS will be discussed by comparison with P450cam and model compounds.

Original languageEnglish
Pages (from-to)5695-5701
Number of pages7
JournalBiochemistry
Volume41
Issue number18
DOIs
StatePublished - 7 May 2002

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Nitric Oxide Synthase Type III
Vibration
Camphor 5-Monooxygenase
Heme
Chloride Peroxidase
Substrates
Hemin
Holoenzymes
Pyrroles
Cytochromes
Stretching
Cysteine
Arginine
Raman scattering
Hydrogen bonds
Proteins
Iron
Tuning
Hydrogen Bonding
Ligands

Cite this

Schelvis, Johannes ; Berka, Vladimir ; Babcock, Gerald T. ; Tsai, Ah Lim. / Resonance Raman detection of the Fe-S bond in endothelial nitric oxide synthase. In: Biochemistry. 2002 ; Vol. 41, No. 18. pp. 5695-5701.
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Resonance Raman detection of the Fe-S bond in endothelial nitric oxide synthase. / Schelvis, Johannes; Berka, Vladimir; Babcock, Gerald T.; Tsai, Ah Lim.

In: Biochemistry, Vol. 41, No. 18, 07.05.2002, p. 5695-5701.

Research output: Contribution to journalArticle

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