Structural and functional insights into lysine acetylation of cytochrome c using mimetic point mutants

Abstract Post‐translational modifications frequently modulate protein functions. Lysine acetylation in particular plays a key role in interactions between respiratory cytochrome c and its metabolic partners. To date, in vivo acetylation of lysines at positions 8 and 53 has specifically been identifi...

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Autores principales:	Inmaculada Márquez, Gonzalo Pérez‐Mejías, Alejandra Guerra‐Castellano, José Luis Olloqui‐Sariego, Rafael Andreu, Juan José Calvente, Miguel A. De la Rosa, Irene Díaz‐Moreno
Formato:	article
Lenguaje:	EN
Publicado:	Wiley 2021
Materias:	acetylation bioenergetics cytochrome c electron transport chain post‐translational modifications Biology (General) QH301-705.5
Acceso en línea:	https://doaj.org/article/f09c544c2ba24db692de7b159c0df6a8
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spelling	oai:doaj.org-article:f09c544c2ba24db692de7b159c0df6a82021-12-01T13:36:12ZStructural and functional insights into lysine acetylation of cytochrome c using mimetic point mutants2211-546310.1002/2211-5463.13284https://doaj.org/article/f09c544c2ba24db692de7b159c0df6a82021-12-01T00:00:00Zhttps://doi.org/10.1002/2211-5463.13284https://doaj.org/toc/2211-5463Abstract Post‐translational modifications frequently modulate protein functions. Lysine acetylation in particular plays a key role in interactions between respiratory cytochrome c and its metabolic partners. To date, in vivo acetylation of lysines at positions 8 and 53 has specifically been identified in mammalian cytochrome c, but little is known about the structural basis of acetylation‐induced functional changes. Here, we independently replaced these two residues in recombinant human cytochrome c with glutamine to mimic lysine acetylation and then characterized the structure and function of the resulting K8Q and K53Q mutants. We found that the physicochemical features were mostly unchanged in the two acetyl‐mimetic mutants, but their thermal stability was significantly altered. NMR chemical shift perturbations of the backbone amide resonances revealed local structural changes, and the thermodynamics and kinetics of electron transfer in mutants immobilized on gold electrodes showed an increase in both protein dynamics and solvent involvement in the redox process. We also observed that the K8Q (but not the K53Q) mutation slightly increased the binding affinity of cytochrome c to its physiological electron donor, cytochrome c1—which is a component of mitochondrial complex III, or cytochrome bc1—thus suggesting that Lys8 (but not Lys53) is located in the interaction area. Finally, the K8Q and K53Q mutants exhibited reduced efficiency as electron donors to complex IV, or cytochrome c oxidase.Inmaculada MárquezGonzalo Pérez‐MejíasAlejandra Guerra‐CastellanoJosé Luis Olloqui‐SariegoRafael AndreuJuan José CalventeMiguel A. De la RosaIrene Díaz‐MorenoWileyarticleacetylationbioenergeticscytochrome celectron transport chainpost‐translational modificationsBiology (General)QH301-705.5ENFEBS Open Bio, Vol 11, Iss 12, Pp 3304-3323 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	acetylation bioenergetics cytochrome c electron transport chain post‐translational modifications Biology (General) QH301-705.5
spellingShingle	acetylation bioenergetics cytochrome c electron transport chain post‐translational modifications Biology (General) QH301-705.5 Inmaculada Márquez Gonzalo Pérez‐Mejías Alejandra Guerra‐Castellano José Luis Olloqui‐Sariego Rafael Andreu Juan José Calvente Miguel A. De la Rosa Irene Díaz‐Moreno Structural and functional insights into lysine acetylation of cytochrome c using mimetic point mutants
description	Abstract Post‐translational modifications frequently modulate protein functions. Lysine acetylation in particular plays a key role in interactions between respiratory cytochrome c and its metabolic partners. To date, in vivo acetylation of lysines at positions 8 and 53 has specifically been identified in mammalian cytochrome c, but little is known about the structural basis of acetylation‐induced functional changes. Here, we independently replaced these two residues in recombinant human cytochrome c with glutamine to mimic lysine acetylation and then characterized the structure and function of the resulting K8Q and K53Q mutants. We found that the physicochemical features were mostly unchanged in the two acetyl‐mimetic mutants, but their thermal stability was significantly altered. NMR chemical shift perturbations of the backbone amide resonances revealed local structural changes, and the thermodynamics and kinetics of electron transfer in mutants immobilized on gold electrodes showed an increase in both protein dynamics and solvent involvement in the redox process. We also observed that the K8Q (but not the K53Q) mutation slightly increased the binding affinity of cytochrome c to its physiological electron donor, cytochrome c1—which is a component of mitochondrial complex III, or cytochrome bc1—thus suggesting that Lys8 (but not Lys53) is located in the interaction area. Finally, the K8Q and K53Q mutants exhibited reduced efficiency as electron donors to complex IV, or cytochrome c oxidase.
format	article
author	Inmaculada Márquez Gonzalo Pérez‐Mejías Alejandra Guerra‐Castellano José Luis Olloqui‐Sariego Rafael Andreu Juan José Calvente Miguel A. De la Rosa Irene Díaz‐Moreno
author_facet	Inmaculada Márquez Gonzalo Pérez‐Mejías Alejandra Guerra‐Castellano José Luis Olloqui‐Sariego Rafael Andreu Juan José Calvente Miguel A. De la Rosa Irene Díaz‐Moreno
author_sort	Inmaculada Márquez
title	Structural and functional insights into lysine acetylation of cytochrome c using mimetic point mutants
title_short	Structural and functional insights into lysine acetylation of cytochrome c using mimetic point mutants
title_full	Structural and functional insights into lysine acetylation of cytochrome c using mimetic point mutants
title_fullStr	Structural and functional insights into lysine acetylation of cytochrome c using mimetic point mutants
title_full_unstemmed	Structural and functional insights into lysine acetylation of cytochrome c using mimetic point mutants
title_sort	structural and functional insights into lysine acetylation of cytochrome c using mimetic point mutants
publisher	Wiley
publishDate	2021
url	https://doaj.org/article/f09c544c2ba24db692de7b159c0df6a8
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_version_	1718405150216814592

Structural and functional insights into lysine acetylation of cytochrome c using mimetic point mutants

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