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Exploring O2 diffusion in A-type cytochrome c oxidases: molecular dynamics simulations uncover two alternative channels towards the binuclear site.

Oliveira AS, Damas JM, Baptista AM, Soares CM - PLoS Comput. Biol. (2014)

Bottom Line: These enzymes couple dioxygen (O2) reduction to the generation of a transmembrane electrochemical proton gradient.In this work, we determined the O2 distribution within Ccox from Rhodobacter sphaeroides, in the fully reduced state, in order to identify and characterize all the putative O2 channels leading towards the BNC.Furthermore, our results show that, in this Ccox, the most likely (energetically preferred) routes for O2 to reach the BNC are the alternative channels, rather than the X-ray inferred pathway.

View Article: PubMed Central - PubMed

Affiliation: ITQB - Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa, Oeiras, Portugal.

ABSTRACT
Cytochrome c oxidases (Ccoxs) are the terminal enzymes of the respiratory chain in mitochondria and most bacteria. These enzymes couple dioxygen (O2) reduction to the generation of a transmembrane electrochemical proton gradient. Despite decades of research and the availability of a large amount of structural and biochemical data available for the A-type Ccox family, little is known about the channel(s) used by O2 to travel from the solvent/membrane to the heme a3-CuB binuclear center (BNC). Moreover, the identification of all possible O2 channels as well as the atomic details of O2 diffusion is essential for the understanding of the working mechanisms of the A-type Ccox. In this work, we determined the O2 distribution within Ccox from Rhodobacter sphaeroides, in the fully reduced state, in order to identify and characterize all the putative O2 channels leading towards the BNC. For that, we use an integrated strategy combining atomistic molecular dynamics (MD) simulations (with and without explicit O2 molecules) and implicit ligand sampling (ILS) calculations. Based on the 3D free energy map for O2 inside Ccox, three channels were identified, all starting in the membrane hydrophobic region and connecting the surface of the protein to the BNC. One of these channels corresponds to the pathway inferred from the X-ray data available, whereas the other two are alternative routes for O2 to reach the BNC. Both alternative O2 channels start in the membrane spanning region and terminate close to Y288I. These channels are a combination of multiple transiently interconnected hydrophobic cavities, whose opening and closure is regulated by the thermal fluctuations of the lining residues. Furthermore, our results show that, in this Ccox, the most likely (energetically preferred) routes for O2 to reach the BNC are the alternative channels, rather than the X-ray inferred pathway.

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Structure of the membrane bound Ccox from R. sphaeroides[15]. For simplicity, only subunits I and II are represented. A) Architecture of the Ccox core functional unit formed by subunits I (colored in green) and II (colored in cyan). The small blue spheres represent the water molecules resolved in the D- and K-pathways in the X-ray structure [15]. During an O2 reduction cycle, the electrons are sequentially donated from cyt c and delivered to the BNC, via CuA and heme a, like in the scheme: cyt c → CuA → heme a → BNC (heme a3/CuB) → O2. B) Putative O2 channel inferred from the X-ray data available. The grey mesh represents the putative O2 channel and it was calculated with the program HOLLOW [92].
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pcbi-1004010-g001: Structure of the membrane bound Ccox from R. sphaeroides[15]. For simplicity, only subunits I and II are represented. A) Architecture of the Ccox core functional unit formed by subunits I (colored in green) and II (colored in cyan). The small blue spheres represent the water molecules resolved in the D- and K-pathways in the X-ray structure [15]. During an O2 reduction cycle, the electrons are sequentially donated from cyt c and delivered to the BNC, via CuA and heme a, like in the scheme: cyt c → CuA → heme a → BNC (heme a3/CuB) → O2. B) Putative O2 channel inferred from the X-ray data available. The grey mesh represents the putative O2 channel and it was calculated with the program HOLLOW [92].

Mentions: Based on structural and phylogenetic analysis, the heme-copper oxidases superfamily is currently divided into three major subfamilies [5]: A, B and C. The main differences between the three families are the pathways and mechanisms of proton transfer/pumping. The A-type Ccoxs, which are the subject of this work, are widespread through all kingdoms of life [5] and among them are the most thoroughly explored Ccoxs [3], [6], such as the bovine heart mitochondria, the Paracoccus (P.) denitrificans and the Rhodobacter (R.) sphaeroides enzymes. These Ccoxs contain, in the catalytic subunit (subunit I), a low spin heme a and a heterodinuclear center named binuclear center, BNC (Fig. 1A). The BNC is deeply buried in the core of the protein and it is formed by a high-spin heme a3 and a copper ion (CuB). In subunit II, these Ccoxs contain only one redox center, a binuclear copper center named CuA, which accepts electrons from the soluble cyt c and then transfers them to the BNC via heme a.


Exploring O2 diffusion in A-type cytochrome c oxidases: molecular dynamics simulations uncover two alternative channels towards the binuclear site.

Oliveira AS, Damas JM, Baptista AM, Soares CM - PLoS Comput. Biol. (2014)

Structure of the membrane bound Ccox from R. sphaeroides[15]. For simplicity, only subunits I and II are represented. A) Architecture of the Ccox core functional unit formed by subunits I (colored in green) and II (colored in cyan). The small blue spheres represent the water molecules resolved in the D- and K-pathways in the X-ray structure [15]. During an O2 reduction cycle, the electrons are sequentially donated from cyt c and delivered to the BNC, via CuA and heme a, like in the scheme: cyt c → CuA → heme a → BNC (heme a3/CuB) → O2. B) Putative O2 channel inferred from the X-ray data available. The grey mesh represents the putative O2 channel and it was calculated with the program HOLLOW [92].
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4256069&req=5

pcbi-1004010-g001: Structure of the membrane bound Ccox from R. sphaeroides[15]. For simplicity, only subunits I and II are represented. A) Architecture of the Ccox core functional unit formed by subunits I (colored in green) and II (colored in cyan). The small blue spheres represent the water molecules resolved in the D- and K-pathways in the X-ray structure [15]. During an O2 reduction cycle, the electrons are sequentially donated from cyt c and delivered to the BNC, via CuA and heme a, like in the scheme: cyt c → CuA → heme a → BNC (heme a3/CuB) → O2. B) Putative O2 channel inferred from the X-ray data available. The grey mesh represents the putative O2 channel and it was calculated with the program HOLLOW [92].
Mentions: Based on structural and phylogenetic analysis, the heme-copper oxidases superfamily is currently divided into three major subfamilies [5]: A, B and C. The main differences between the three families are the pathways and mechanisms of proton transfer/pumping. The A-type Ccoxs, which are the subject of this work, are widespread through all kingdoms of life [5] and among them are the most thoroughly explored Ccoxs [3], [6], such as the bovine heart mitochondria, the Paracoccus (P.) denitrificans and the Rhodobacter (R.) sphaeroides enzymes. These Ccoxs contain, in the catalytic subunit (subunit I), a low spin heme a and a heterodinuclear center named binuclear center, BNC (Fig. 1A). The BNC is deeply buried in the core of the protein and it is formed by a high-spin heme a3 and a copper ion (CuB). In subunit II, these Ccoxs contain only one redox center, a binuclear copper center named CuA, which accepts electrons from the soluble cyt c and then transfers them to the BNC via heme a.

Bottom Line: These enzymes couple dioxygen (O2) reduction to the generation of a transmembrane electrochemical proton gradient.In this work, we determined the O2 distribution within Ccox from Rhodobacter sphaeroides, in the fully reduced state, in order to identify and characterize all the putative O2 channels leading towards the BNC.Furthermore, our results show that, in this Ccox, the most likely (energetically preferred) routes for O2 to reach the BNC are the alternative channels, rather than the X-ray inferred pathway.

View Article: PubMed Central - PubMed

Affiliation: ITQB - Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa, Oeiras, Portugal.

ABSTRACT
Cytochrome c oxidases (Ccoxs) are the terminal enzymes of the respiratory chain in mitochondria and most bacteria. These enzymes couple dioxygen (O2) reduction to the generation of a transmembrane electrochemical proton gradient. Despite decades of research and the availability of a large amount of structural and biochemical data available for the A-type Ccox family, little is known about the channel(s) used by O2 to travel from the solvent/membrane to the heme a3-CuB binuclear center (BNC). Moreover, the identification of all possible O2 channels as well as the atomic details of O2 diffusion is essential for the understanding of the working mechanisms of the A-type Ccox. In this work, we determined the O2 distribution within Ccox from Rhodobacter sphaeroides, in the fully reduced state, in order to identify and characterize all the putative O2 channels leading towards the BNC. For that, we use an integrated strategy combining atomistic molecular dynamics (MD) simulations (with and without explicit O2 molecules) and implicit ligand sampling (ILS) calculations. Based on the 3D free energy map for O2 inside Ccox, three channels were identified, all starting in the membrane hydrophobic region and connecting the surface of the protein to the BNC. One of these channels corresponds to the pathway inferred from the X-ray data available, whereas the other two are alternative routes for O2 to reach the BNC. Both alternative O2 channels start in the membrane spanning region and terminate close to Y288I. These channels are a combination of multiple transiently interconnected hydrophobic cavities, whose opening and closure is regulated by the thermal fluctuations of the lining residues. Furthermore, our results show that, in this Ccox, the most likely (energetically preferred) routes for O2 to reach the BNC are the alternative channels, rather than the X-ray inferred pathway.

Show MeSH
Related in: MedlinePlus