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Structural and spectroscopic characterisation of a heme peroxidase from sorghum
A cationic class III peroxidase from Sorghum bicolor was purified to homogeneity. The enzyme contains a high-spin heme, as evidenced by UV–visible spectroscopy and EPR. Steady state oxidation of guaiacol was demonstrated and the enzyme was shown to have higher activity in the presence of calcium ion...
Autores principales: | , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Springer Berlin Heidelberg
2015
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4771821/ https://www.ncbi.nlm.nih.gov/pubmed/26666777 http://dx.doi.org/10.1007/s00775-015-1313-z |
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author | Nnamchi, Chukwudi I. Parkin, Gary Efimov, Igor Basran, Jaswir Kwon, Hanna Svistunenko, Dimitri A. Agirre, Jon Okolo, Bartholomew N. Moneke, Anene Nwanguma, Bennett C. Moody, Peter C. E. Raven, Emma L. |
author_facet | Nnamchi, Chukwudi I. Parkin, Gary Efimov, Igor Basran, Jaswir Kwon, Hanna Svistunenko, Dimitri A. Agirre, Jon Okolo, Bartholomew N. Moneke, Anene Nwanguma, Bennett C. Moody, Peter C. E. Raven, Emma L. |
author_sort | Nnamchi, Chukwudi I. |
collection | PubMed |
description | A cationic class III peroxidase from Sorghum bicolor was purified to homogeneity. The enzyme contains a high-spin heme, as evidenced by UV–visible spectroscopy and EPR. Steady state oxidation of guaiacol was demonstrated and the enzyme was shown to have higher activity in the presence of calcium ions. A Fe(III)/Fe(II) reduction potential of −266 mV vs NHE was determined. Stopped-flow experiments with H(2)O(2) showed formation of a typical peroxidase Compound I species, which converts to Compound II in the presence of calcium. A crystal structure of the enzyme is reported, the first for a sorghum peroxidase. The structure reveals an active site that is analogous to those for other class I heme peroxidase, and a substrate binding site (assigned as arising from binding of indole-3-acetic acid) at the γ-heme edge. Metal binding sites are observed in the structure on the distal (assigned as a Na(+) ion) and proximal (assigned as a Ca(2+)) sides of the heme, which is consistent with the Ca(2+)-dependence of the steady state and pre-steady state kinetics. It is probably the case that the structural integrity (and, thus, the catalytic activity) of the sorghum enzyme is dependent on metal ion incorporation at these positions. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s00775-015-1313-z) contains supplementary material, which is available to authorized users. |
format | Online Article Text |
id | pubmed-4771821 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2015 |
publisher | Springer Berlin Heidelberg |
record_format | MEDLINE/PubMed |
spelling | pubmed-47718212016-03-22 Structural and spectroscopic characterisation of a heme peroxidase from sorghum Nnamchi, Chukwudi I. Parkin, Gary Efimov, Igor Basran, Jaswir Kwon, Hanna Svistunenko, Dimitri A. Agirre, Jon Okolo, Bartholomew N. Moneke, Anene Nwanguma, Bennett C. Moody, Peter C. E. Raven, Emma L. J Biol Inorg Chem Original Paper A cationic class III peroxidase from Sorghum bicolor was purified to homogeneity. The enzyme contains a high-spin heme, as evidenced by UV–visible spectroscopy and EPR. Steady state oxidation of guaiacol was demonstrated and the enzyme was shown to have higher activity in the presence of calcium ions. A Fe(III)/Fe(II) reduction potential of −266 mV vs NHE was determined. Stopped-flow experiments with H(2)O(2) showed formation of a typical peroxidase Compound I species, which converts to Compound II in the presence of calcium. A crystal structure of the enzyme is reported, the first for a sorghum peroxidase. The structure reveals an active site that is analogous to those for other class I heme peroxidase, and a substrate binding site (assigned as arising from binding of indole-3-acetic acid) at the γ-heme edge. Metal binding sites are observed in the structure on the distal (assigned as a Na(+) ion) and proximal (assigned as a Ca(2+)) sides of the heme, which is consistent with the Ca(2+)-dependence of the steady state and pre-steady state kinetics. It is probably the case that the structural integrity (and, thus, the catalytic activity) of the sorghum enzyme is dependent on metal ion incorporation at these positions. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s00775-015-1313-z) contains supplementary material, which is available to authorized users. Springer Berlin Heidelberg 2015-12-14 2016 /pmc/articles/PMC4771821/ /pubmed/26666777 http://dx.doi.org/10.1007/s00775-015-1313-z Text en © The Author(s) 2015 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. |
spellingShingle | Original Paper Nnamchi, Chukwudi I. Parkin, Gary Efimov, Igor Basran, Jaswir Kwon, Hanna Svistunenko, Dimitri A. Agirre, Jon Okolo, Bartholomew N. Moneke, Anene Nwanguma, Bennett C. Moody, Peter C. E. Raven, Emma L. Structural and spectroscopic characterisation of a heme peroxidase from sorghum |
title | Structural and spectroscopic characterisation of a heme peroxidase from sorghum |
title_full | Structural and spectroscopic characterisation of a heme peroxidase from sorghum |
title_fullStr | Structural and spectroscopic characterisation of a heme peroxidase from sorghum |
title_full_unstemmed | Structural and spectroscopic characterisation of a heme peroxidase from sorghum |
title_short | Structural and spectroscopic characterisation of a heme peroxidase from sorghum |
title_sort | structural and spectroscopic characterisation of a heme peroxidase from sorghum |
topic | Original Paper |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4771821/ https://www.ncbi.nlm.nih.gov/pubmed/26666777 http://dx.doi.org/10.1007/s00775-015-1313-z |
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