Direct binding of hydroxylamine to the heme iron of Arthromyces ramosus peroxidase. Substrate analogue that inhibits compound I formation in a competitive manner

H. Wariishi, D. Nonaka, T. Johjima, N. Nakamura, Y. Naruta, S. Kubo, K. Fukuyama

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

The interaction of hydroxylamine (HA) with Arthromyces ramosus peroxidase (ARP) was investigated by kinetic, spectroscopic, and x-ray crystallographic techniques. HA inhibited the reaction of native ARP with H2O2 in a competitive manner. Electron absorption and resonance Raman spectroscopic studies indicated that pentacoordinate high spin species of native ARP are converted to hexacoordinate low spin species upon the addition of HA, strongly suggesting the occurrence of a direct interaction of HA with ARP heine iron. Kinetic analysis exhibited that the apparent dissociation constant is 6.2 mM at pH 7.0 and that only one HA molecule likely binds to the vicinity of the heme. pH dependence of HA binding suggested that the nitrogen atom of HA could be involved in the interaction with the heine iron. X-ray crystallographic analysis of ARP in complex with HA at 2.0 Å resolution revealed that the electron density ascribed to HA is located in the distal pocket between the heine iron and the distal His56. HA seems to directly interact with the heine iron but is too far away to interact with Arg52. In HA, it is likely that the nitrogen atom is coordinated to the heine iron and that hydroxyl group is hydrogen bonded to the distal His56.

Original languageEnglish
Pages (from-to)32919-32924
Number of pages6
JournalJournal of Biological Chemistry
Volume275
Issue number42
DOIs
Publication statusPublished - Oct 20 2000
Externally publishedYes

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Hydroxylamine
Heme
Peroxidase
Iron
Substrates
Nitrogen
Electron absorption
Electron resonance
X-Rays
Electrons
X rays
Atoms
Kinetics
Hydroxyl Radical
Carrier concentration
Hydrogen

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Direct binding of hydroxylamine to the heme iron of Arthromyces ramosus peroxidase. Substrate analogue that inhibits compound I formation in a competitive manner. / Wariishi, H.; Nonaka, D.; Johjima, T.; Nakamura, N.; Naruta, Y.; Kubo, S.; Fukuyama, K.

In: Journal of Biological Chemistry, Vol. 275, No. 42, 20.10.2000, p. 32919-32924.

Research output: Contribution to journalArticle

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AU - Nonaka, D.

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AU - Nakamura, N.

AU - Naruta, Y.

AU - Kubo, S.

AU - Fukuyama, K.

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