Isolation and characterization of a microperoxidase-8 with a modified histidine axial ligand

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Abstract

Microperoxidase-8, Fe(III)MP-8, the heme octapeptide obtained by horse heart cytochrome c digestion, was studied in the presence of H 2O 2. A modified form of the catalyst was isolated by HPLC and showed a UV/visible spectrum similar to that of Fe(III)MP-8. ESI-MS measurements revealed a 16 Da increase in molecular mass for the modified catalyst when compared to Fe(III)MP-8, suggesting the insertion of an oxygen atom. ESI-MS 2 fragmentation measurements point at oxygen incorporation on the His18 residue of the octapeptide of the modified catalyst. Comparison of the 1H NMR chemical shifts of the methyl protons of the porphyrin ring of Fe(III)MP-8 and the modified catalyst shows a large shift for especially the 3-methyl and 5-methyl resonances, whereas the other 1H NMR chemical shifts are almost unaffected. These observations can best be ascribed to a reorientation of the histidine axial ligand. The latter is suggested to be the consequence of an oxygen insertion, possibly on the imidazole ring of His18, thereby corroborating the data obtained by ESI-MS 2. 1H NMR NOE difference measurements on Fe(III)MP-8 and on the modified catalyst supported the assignment of the Hd2 and He1 protons of the His18 imidazole ring. The ring amine proton Hd1 could not be detected in both forms of the catalyst. For Fe(III)MP-8 this absence of the Hd1 resonance can be ascribed to fast H/D exchange. For the modified catalyst the NMR data are not contradictory, with an oxygen insertion on position d1 of the His18 imidazole ring with a fast H/D exchanging hydroxyl proton. Together these data converge in suggesting the H 2O 2 modified catalyst bears a hydroxylated His18 axial ligand. The mechanism that could underlie Fe(III)MP-8 axial histidine hydroxylation is further discussed.
LanguageEnglish
Pages870-878
JournalJournal of Biological Inorganic Chemistry
Volume7
Issue number7-8
DOIs
Publication statusPublished - 2002

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Histidine
Protons
Oxygen
Ligands
Catalysts
Nuclear magnetic resonance
Chemical shift
Porphyrins
Hydroxylation
Cytochromes c
Heme
Hydroxyl Radical
Horses
Amines
Digestion
High Pressure Liquid Chromatography
Molecular mass
microperoxidase
Catalyst supports
Proton Magnetic Resonance Spectroscopy

Cite this

@article{e0819536f82d4e9386a39e1ea170ab98,
title = "Isolation and characterization of a microperoxidase-8 with a modified histidine axial ligand",
abstract = "Microperoxidase-8, Fe(III)MP-8, the heme octapeptide obtained by horse heart cytochrome c digestion, was studied in the presence of H 2O 2. A modified form of the catalyst was isolated by HPLC and showed a UV/visible spectrum similar to that of Fe(III)MP-8. ESI-MS measurements revealed a 16 Da increase in molecular mass for the modified catalyst when compared to Fe(III)MP-8, suggesting the insertion of an oxygen atom. ESI-MS 2 fragmentation measurements point at oxygen incorporation on the His18 residue of the octapeptide of the modified catalyst. Comparison of the 1H NMR chemical shifts of the methyl protons of the porphyrin ring of Fe(III)MP-8 and the modified catalyst shows a large shift for especially the 3-methyl and 5-methyl resonances, whereas the other 1H NMR chemical shifts are almost unaffected. These observations can best be ascribed to a reorientation of the histidine axial ligand. The latter is suggested to be the consequence of an oxygen insertion, possibly on the imidazole ring of His18, thereby corroborating the data obtained by ESI-MS 2. 1H NMR NOE difference measurements on Fe(III)MP-8 and on the modified catalyst supported the assignment of the Hd2 and He1 protons of the His18 imidazole ring. The ring amine proton Hd1 could not be detected in both forms of the catalyst. For Fe(III)MP-8 this absence of the Hd1 resonance can be ascribed to fast H/D exchange. For the modified catalyst the NMR data are not contradictory, with an oxygen insertion on position d1 of the His18 imidazole ring with a fast H/D exchanging hydroxyl proton. Together these data converge in suggesting the H 2O 2 modified catalyst bears a hydroxylated His18 axial ligand. The mechanism that could underlie Fe(III)MP-8 axial histidine hydroxylation is further discussed.",
author = "J.L. Primus and S. Boeren and M.W.F. Nielen and J. Vervoort and L. Banci and I.M.C.M. Rietjens",
year = "2002",
doi = "10.1007/s00775-002-0373-z",
language = "English",
volume = "7",
pages = "870--878",
journal = "Journal of Biological Inorganic Chemistry",
issn = "0949-8257",
publisher = "Springer Verlag",
number = "7-8",

}

Isolation and characterization of a microperoxidase-8 with a modified histidine axial ligand. / Primus, J.L.; Boeren, S.; Nielen, M.W.F.; Vervoort, J.; Banci, L.; Rietjens, I.M.C.M.

In: Journal of Biological Inorganic Chemistry, Vol. 7, No. 7-8, 2002, p. 870-878.

Research output: Contribution to journalArticleAcademicpeer-review

TY - JOUR

T1 - Isolation and characterization of a microperoxidase-8 with a modified histidine axial ligand

AU - Primus, J.L.

AU - Boeren, S.

AU - Nielen, M.W.F.

AU - Vervoort, J.

AU - Banci, L.

AU - Rietjens, I.M.C.M.

PY - 2002

Y1 - 2002

N2 - Microperoxidase-8, Fe(III)MP-8, the heme octapeptide obtained by horse heart cytochrome c digestion, was studied in the presence of H 2O 2. A modified form of the catalyst was isolated by HPLC and showed a UV/visible spectrum similar to that of Fe(III)MP-8. ESI-MS measurements revealed a 16 Da increase in molecular mass for the modified catalyst when compared to Fe(III)MP-8, suggesting the insertion of an oxygen atom. ESI-MS 2 fragmentation measurements point at oxygen incorporation on the His18 residue of the octapeptide of the modified catalyst. Comparison of the 1H NMR chemical shifts of the methyl protons of the porphyrin ring of Fe(III)MP-8 and the modified catalyst shows a large shift for especially the 3-methyl and 5-methyl resonances, whereas the other 1H NMR chemical shifts are almost unaffected. These observations can best be ascribed to a reorientation of the histidine axial ligand. The latter is suggested to be the consequence of an oxygen insertion, possibly on the imidazole ring of His18, thereby corroborating the data obtained by ESI-MS 2. 1H NMR NOE difference measurements on Fe(III)MP-8 and on the modified catalyst supported the assignment of the Hd2 and He1 protons of the His18 imidazole ring. The ring amine proton Hd1 could not be detected in both forms of the catalyst. For Fe(III)MP-8 this absence of the Hd1 resonance can be ascribed to fast H/D exchange. For the modified catalyst the NMR data are not contradictory, with an oxygen insertion on position d1 of the His18 imidazole ring with a fast H/D exchanging hydroxyl proton. Together these data converge in suggesting the H 2O 2 modified catalyst bears a hydroxylated His18 axial ligand. The mechanism that could underlie Fe(III)MP-8 axial histidine hydroxylation is further discussed.

AB - Microperoxidase-8, Fe(III)MP-8, the heme octapeptide obtained by horse heart cytochrome c digestion, was studied in the presence of H 2O 2. A modified form of the catalyst was isolated by HPLC and showed a UV/visible spectrum similar to that of Fe(III)MP-8. ESI-MS measurements revealed a 16 Da increase in molecular mass for the modified catalyst when compared to Fe(III)MP-8, suggesting the insertion of an oxygen atom. ESI-MS 2 fragmentation measurements point at oxygen incorporation on the His18 residue of the octapeptide of the modified catalyst. Comparison of the 1H NMR chemical shifts of the methyl protons of the porphyrin ring of Fe(III)MP-8 and the modified catalyst shows a large shift for especially the 3-methyl and 5-methyl resonances, whereas the other 1H NMR chemical shifts are almost unaffected. These observations can best be ascribed to a reorientation of the histidine axial ligand. The latter is suggested to be the consequence of an oxygen insertion, possibly on the imidazole ring of His18, thereby corroborating the data obtained by ESI-MS 2. 1H NMR NOE difference measurements on Fe(III)MP-8 and on the modified catalyst supported the assignment of the Hd2 and He1 protons of the His18 imidazole ring. The ring amine proton Hd1 could not be detected in both forms of the catalyst. For Fe(III)MP-8 this absence of the Hd1 resonance can be ascribed to fast H/D exchange. For the modified catalyst the NMR data are not contradictory, with an oxygen insertion on position d1 of the His18 imidazole ring with a fast H/D exchanging hydroxyl proton. Together these data converge in suggesting the H 2O 2 modified catalyst bears a hydroxylated His18 axial ligand. The mechanism that could underlie Fe(III)MP-8 axial histidine hydroxylation is further discussed.

U2 - 10.1007/s00775-002-0373-z

DO - 10.1007/s00775-002-0373-z

M3 - Article

VL - 7

SP - 870

EP - 878

JO - Journal of Biological Inorganic Chemistry

T2 - Journal of Biological Inorganic Chemistry

JF - Journal of Biological Inorganic Chemistry

SN - 0949-8257

IS - 7-8

ER -