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Multiple Mutations in the Non-Ordered Red Ω-Loop Enhance the Membrane-Permeabilizing and Peroxidase-like Activity of Cytochrome c

A key event in the cytochrome c-dependent apoptotic pathway is the permeabilization of the outer mitochondrial membrane, resulting in the release of various apoptogenic factors, including cytochrome c, into the cytosol. It is believed that the permeabilization of the outer mitochondrial membrane can...

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Autores principales: Chertkova, Rita V., Firsov, Alexander M., Brazhe, Nadezda A., Nikelshparg, Evelina I., Bochkova, Zhanna V., Bryantseva, Tatyana V., Semenova, Marina A., Baizhumanov, Adil A., Kotova, Elena A., Kirpichnikov, Mikhail P., Maksimov, Georgy V., Antonenko, Yuriy N., Dolgikh, Dmitry A.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9138828/
https://www.ncbi.nlm.nih.gov/pubmed/35625593
http://dx.doi.org/10.3390/biom12050665
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author Chertkova, Rita V.
Firsov, Alexander M.
Brazhe, Nadezda A.
Nikelshparg, Evelina I.
Bochkova, Zhanna V.
Bryantseva, Tatyana V.
Semenova, Marina A.
Baizhumanov, Adil A.
Kotova, Elena A.
Kirpichnikov, Mikhail P.
Maksimov, Georgy V.
Antonenko, Yuriy N.
Dolgikh, Dmitry A.
author_facet Chertkova, Rita V.
Firsov, Alexander M.
Brazhe, Nadezda A.
Nikelshparg, Evelina I.
Bochkova, Zhanna V.
Bryantseva, Tatyana V.
Semenova, Marina A.
Baizhumanov, Adil A.
Kotova, Elena A.
Kirpichnikov, Mikhail P.
Maksimov, Georgy V.
Antonenko, Yuriy N.
Dolgikh, Dmitry A.
author_sort Chertkova, Rita V.
collection PubMed
description A key event in the cytochrome c-dependent apoptotic pathway is the permeabilization of the outer mitochondrial membrane, resulting in the release of various apoptogenic factors, including cytochrome c, into the cytosol. It is believed that the permeabilization of the outer mitochondrial membrane can be induced by the peroxidase activity of cytochrome c in a complex with cardiolipin. Using a number of mutant variants of cytochrome c, we showed that both substitutions of Lys residues from the universal binding site for oppositely charged Glu residues and mutations leading to a decrease in the conformational mobility of the red Ω-loop in almost all cases did not affect the ability of cytochrome c to bind to cardiolipin. At the same time, the peroxidase activity of all mutant variants in a complex with cardiolipin was three to five times higher than that of the wild type. A pronounced increase in the ability to permeabilize the lipid membrane in the presence of hydrogen peroxide, as measured by calcein leakage from liposomes, was observed only in the case of four substitutions in the red Ω-loop (M4 mutant). According to resonance and surface-enhanced Raman spectroscopy, the mutations caused significant changes in the heme of oxidized cytochrome c molecules resulting in an increased probability of the plane heme conformation and the enhancement of the rigidity of the protein surrounding the heme. The binding of wild-type and mutant forms of oxidized cytochrome c to cardiolipin-containing liposomes caused the disordering of the acyl lipid chains that was more pronounced for the M4 mutant. Our findings indicate that the Ω-loop is important for the pore formation in cardiolipin-containing membranes.
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spelling pubmed-91388282022-05-28 Multiple Mutations in the Non-Ordered Red Ω-Loop Enhance the Membrane-Permeabilizing and Peroxidase-like Activity of Cytochrome c Chertkova, Rita V. Firsov, Alexander M. Brazhe, Nadezda A. Nikelshparg, Evelina I. Bochkova, Zhanna V. Bryantseva, Tatyana V. Semenova, Marina A. Baizhumanov, Adil A. Kotova, Elena A. Kirpichnikov, Mikhail P. Maksimov, Georgy V. Antonenko, Yuriy N. Dolgikh, Dmitry A. Biomolecules Article A key event in the cytochrome c-dependent apoptotic pathway is the permeabilization of the outer mitochondrial membrane, resulting in the release of various apoptogenic factors, including cytochrome c, into the cytosol. It is believed that the permeabilization of the outer mitochondrial membrane can be induced by the peroxidase activity of cytochrome c in a complex with cardiolipin. Using a number of mutant variants of cytochrome c, we showed that both substitutions of Lys residues from the universal binding site for oppositely charged Glu residues and mutations leading to a decrease in the conformational mobility of the red Ω-loop in almost all cases did not affect the ability of cytochrome c to bind to cardiolipin. At the same time, the peroxidase activity of all mutant variants in a complex with cardiolipin was three to five times higher than that of the wild type. A pronounced increase in the ability to permeabilize the lipid membrane in the presence of hydrogen peroxide, as measured by calcein leakage from liposomes, was observed only in the case of four substitutions in the red Ω-loop (M4 mutant). According to resonance and surface-enhanced Raman spectroscopy, the mutations caused significant changes in the heme of oxidized cytochrome c molecules resulting in an increased probability of the plane heme conformation and the enhancement of the rigidity of the protein surrounding the heme. The binding of wild-type and mutant forms of oxidized cytochrome c to cardiolipin-containing liposomes caused the disordering of the acyl lipid chains that was more pronounced for the M4 mutant. Our findings indicate that the Ω-loop is important for the pore formation in cardiolipin-containing membranes. MDPI 2022-05-04 /pmc/articles/PMC9138828/ /pubmed/35625593 http://dx.doi.org/10.3390/biom12050665 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/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 (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Chertkova, Rita V.
Firsov, Alexander M.
Brazhe, Nadezda A.
Nikelshparg, Evelina I.
Bochkova, Zhanna V.
Bryantseva, Tatyana V.
Semenova, Marina A.
Baizhumanov, Adil A.
Kotova, Elena A.
Kirpichnikov, Mikhail P.
Maksimov, Georgy V.
Antonenko, Yuriy N.
Dolgikh, Dmitry A.
Multiple Mutations in the Non-Ordered Red Ω-Loop Enhance the Membrane-Permeabilizing and Peroxidase-like Activity of Cytochrome c
title Multiple Mutations in the Non-Ordered Red Ω-Loop Enhance the Membrane-Permeabilizing and Peroxidase-like Activity of Cytochrome c
title_full Multiple Mutations in the Non-Ordered Red Ω-Loop Enhance the Membrane-Permeabilizing and Peroxidase-like Activity of Cytochrome c
title_fullStr Multiple Mutations in the Non-Ordered Red Ω-Loop Enhance the Membrane-Permeabilizing and Peroxidase-like Activity of Cytochrome c
title_full_unstemmed Multiple Mutations in the Non-Ordered Red Ω-Loop Enhance the Membrane-Permeabilizing and Peroxidase-like Activity of Cytochrome c
title_short Multiple Mutations in the Non-Ordered Red Ω-Loop Enhance the Membrane-Permeabilizing and Peroxidase-like Activity of Cytochrome c
title_sort multiple mutations in the non-ordered red ω-loop enhance the membrane-permeabilizing and peroxidase-like activity of cytochrome c
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9138828/
https://www.ncbi.nlm.nih.gov/pubmed/35625593
http://dx.doi.org/10.3390/biom12050665
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