Cargando…

The Reaction of Oxy Hemoglobin with Nitrite: Mechanism, Antioxidant-Modulated Effect, and Implications for Blood Substitute Evaluation

The autocatalytic reaction between nitrite and the oxy form of globins involves free radicals. For myoglobin (Mb), an initial binding of nitrite to the iron-coordinated oxygen molecule was proposed; the resulting ferrous-peroxynitrate species was not detected, but its decay product, the high-valent...

Descripción completa

Detalles Bibliográficos
Autores principales: Hathazi, Denisa, Scurtu, Florina, Bischin, Cristina, Mot, Augustin, Attia, Amr A. A., Kongsted, Jacob, Silaghi-Dumitrescu, Radu
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6017026/
https://www.ncbi.nlm.nih.gov/pubmed/29414908
http://dx.doi.org/10.3390/molecules23020350
_version_ 1783334657163526144
author Hathazi, Denisa
Scurtu, Florina
Bischin, Cristina
Mot, Augustin
Attia, Amr A. A.
Kongsted, Jacob
Silaghi-Dumitrescu, Radu
author_facet Hathazi, Denisa
Scurtu, Florina
Bischin, Cristina
Mot, Augustin
Attia, Amr A. A.
Kongsted, Jacob
Silaghi-Dumitrescu, Radu
author_sort Hathazi, Denisa
collection PubMed
description The autocatalytic reaction between nitrite and the oxy form of globins involves free radicals. For myoglobin (Mb), an initial binding of nitrite to the iron-coordinated oxygen molecule was proposed; the resulting ferrous-peroxynitrate species was not detected, but its decay product, the high-valent ferryl form, was demonstrated in stopped-flow experiments. Reported here are the stopped flow spectra recorded upon mixing oxy Hb (native, as well as chemically-derivatized in the form of several candidates of blood substitutes) with a supraphysiological concentration of nitrite. The data may be fitted to a simple kinetic model involving a transient met-aqua form, in contrast to the ferryl detected in the case of Mb in a similar reaction sequence. These data are in line with a previous observation of a transient accumulation of ferryl Hb under auto-catalytic conditions at much lower concentrations of nitrite (Grubina, R. et al. J. Biol. Chem. 2007, 282, 12916). The simple model for fitting the stopped-flow data leaves a small part of the absorbance changes unaccounted for, unless a fourth species is invoked displaying features similar to the oxy and tentatively assigned as ferrous-peroxynitrate. Density functional theory (DFT) calculations support this latter assignment. The reaction allows for differentiating between the reactivities of various chemically modified hemoglobins, including candidates for blood substitutes. Polymerization of hemoglobin slows the nitrite-induced oxidation, in sharp contrast to oxidative-stress type reactions which are generally accelerated, not inhibited. Sheep hemoglobin is found to be distinctly more resistant to reaction with nitrite compared to bovine Hb, at large nitrite concentrations (stopped-flow experiments directly observing the oxy + nitrite reaction) as well as under auto-catalytic conditions. Copolymerization of Hb with bovine serum albumin (BSA) using glutaraldehyde leads to a distinct increase of the lag time compared to native Hb as well as to any other form of derivatization examined in the present study. The Hb-BSA copolymer also displays a slower initial reaction with nitrite under stopped-flow conditions, compared to native Hb.
format Online
Article
Text
id pubmed-6017026
institution National Center for Biotechnology Information
language English
publishDate 2018
publisher MDPI
record_format MEDLINE/PubMed
spelling pubmed-60170262018-11-13 The Reaction of Oxy Hemoglobin with Nitrite: Mechanism, Antioxidant-Modulated Effect, and Implications for Blood Substitute Evaluation Hathazi, Denisa Scurtu, Florina Bischin, Cristina Mot, Augustin Attia, Amr A. A. Kongsted, Jacob Silaghi-Dumitrescu, Radu Molecules Article The autocatalytic reaction between nitrite and the oxy form of globins involves free radicals. For myoglobin (Mb), an initial binding of nitrite to the iron-coordinated oxygen molecule was proposed; the resulting ferrous-peroxynitrate species was not detected, but its decay product, the high-valent ferryl form, was demonstrated in stopped-flow experiments. Reported here are the stopped flow spectra recorded upon mixing oxy Hb (native, as well as chemically-derivatized in the form of several candidates of blood substitutes) with a supraphysiological concentration of nitrite. The data may be fitted to a simple kinetic model involving a transient met-aqua form, in contrast to the ferryl detected in the case of Mb in a similar reaction sequence. These data are in line with a previous observation of a transient accumulation of ferryl Hb under auto-catalytic conditions at much lower concentrations of nitrite (Grubina, R. et al. J. Biol. Chem. 2007, 282, 12916). The simple model for fitting the stopped-flow data leaves a small part of the absorbance changes unaccounted for, unless a fourth species is invoked displaying features similar to the oxy and tentatively assigned as ferrous-peroxynitrate. Density functional theory (DFT) calculations support this latter assignment. The reaction allows for differentiating between the reactivities of various chemically modified hemoglobins, including candidates for blood substitutes. Polymerization of hemoglobin slows the nitrite-induced oxidation, in sharp contrast to oxidative-stress type reactions which are generally accelerated, not inhibited. Sheep hemoglobin is found to be distinctly more resistant to reaction with nitrite compared to bovine Hb, at large nitrite concentrations (stopped-flow experiments directly observing the oxy + nitrite reaction) as well as under auto-catalytic conditions. Copolymerization of Hb with bovine serum albumin (BSA) using glutaraldehyde leads to a distinct increase of the lag time compared to native Hb as well as to any other form of derivatization examined in the present study. The Hb-BSA copolymer also displays a slower initial reaction with nitrite under stopped-flow conditions, compared to native Hb. MDPI 2018-02-07 /pmc/articles/PMC6017026/ /pubmed/29414908 http://dx.doi.org/10.3390/molecules23020350 Text en © 2018 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Hathazi, Denisa
Scurtu, Florina
Bischin, Cristina
Mot, Augustin
Attia, Amr A. A.
Kongsted, Jacob
Silaghi-Dumitrescu, Radu
The Reaction of Oxy Hemoglobin with Nitrite: Mechanism, Antioxidant-Modulated Effect, and Implications for Blood Substitute Evaluation
title The Reaction of Oxy Hemoglobin with Nitrite: Mechanism, Antioxidant-Modulated Effect, and Implications for Blood Substitute Evaluation
title_full The Reaction of Oxy Hemoglobin with Nitrite: Mechanism, Antioxidant-Modulated Effect, and Implications for Blood Substitute Evaluation
title_fullStr The Reaction of Oxy Hemoglobin with Nitrite: Mechanism, Antioxidant-Modulated Effect, and Implications for Blood Substitute Evaluation
title_full_unstemmed The Reaction of Oxy Hemoglobin with Nitrite: Mechanism, Antioxidant-Modulated Effect, and Implications for Blood Substitute Evaluation
title_short The Reaction of Oxy Hemoglobin with Nitrite: Mechanism, Antioxidant-Modulated Effect, and Implications for Blood Substitute Evaluation
title_sort reaction of oxy hemoglobin with nitrite: mechanism, antioxidant-modulated effect, and implications for blood substitute evaluation
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6017026/
https://www.ncbi.nlm.nih.gov/pubmed/29414908
http://dx.doi.org/10.3390/molecules23020350
work_keys_str_mv AT hathazidenisa thereactionofoxyhemoglobinwithnitritemechanismantioxidantmodulatedeffectandimplicationsforbloodsubstituteevaluation
AT scurtuflorina thereactionofoxyhemoglobinwithnitritemechanismantioxidantmodulatedeffectandimplicationsforbloodsubstituteevaluation
AT bischincristina thereactionofoxyhemoglobinwithnitritemechanismantioxidantmodulatedeffectandimplicationsforbloodsubstituteevaluation
AT motaugustin thereactionofoxyhemoglobinwithnitritemechanismantioxidantmodulatedeffectandimplicationsforbloodsubstituteevaluation
AT attiaamraa thereactionofoxyhemoglobinwithnitritemechanismantioxidantmodulatedeffectandimplicationsforbloodsubstituteevaluation
AT kongstedjacob thereactionofoxyhemoglobinwithnitritemechanismantioxidantmodulatedeffectandimplicationsforbloodsubstituteevaluation
AT silaghidumitrescuradu thereactionofoxyhemoglobinwithnitritemechanismantioxidantmodulatedeffectandimplicationsforbloodsubstituteevaluation
AT hathazidenisa reactionofoxyhemoglobinwithnitritemechanismantioxidantmodulatedeffectandimplicationsforbloodsubstituteevaluation
AT scurtuflorina reactionofoxyhemoglobinwithnitritemechanismantioxidantmodulatedeffectandimplicationsforbloodsubstituteevaluation
AT bischincristina reactionofoxyhemoglobinwithnitritemechanismantioxidantmodulatedeffectandimplicationsforbloodsubstituteevaluation
AT motaugustin reactionofoxyhemoglobinwithnitritemechanismantioxidantmodulatedeffectandimplicationsforbloodsubstituteevaluation
AT attiaamraa reactionofoxyhemoglobinwithnitritemechanismantioxidantmodulatedeffectandimplicationsforbloodsubstituteevaluation
AT kongstedjacob reactionofoxyhemoglobinwithnitritemechanismantioxidantmodulatedeffectandimplicationsforbloodsubstituteevaluation
AT silaghidumitrescuradu reactionofoxyhemoglobinwithnitritemechanismantioxidantmodulatedeffectandimplicationsforbloodsubstituteevaluation