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Lipid-Loving ANTs: Molecular Simulations of CardiolipinInteractions and the Organization of the Adenine Nucleotide Translocase in Model Mitochondrial Membranes

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ABSTRACT

The exchange of ADPand ATP across the inner mitochondrial membraneis a fundamental cellular process. This exchange is facilitated bythe adenine nucleotide translocase, the structure and function ofwhich are critically dependent on the signature phospholipid of mitochondria,cardiolipin (CL). Here we employ multiscale molecular dynamics simulationsto investigate CL interactions within a membrane environment. Usingsimulations at both coarse-grained and atomistic resolutions, we identifythree CL binding sites on the translocase, in agreement with thoseseen in crystal structures and inferred from nuclear magnetic resonancemeasurements. Characterization of the free energy landscape for laterallipid interaction via potential of mean force calculations demonstratesthe strength of interaction compared to those of binding sites onother mitochondrial membrane proteins, as well as their selectivityfor CL over other phospholipids. Extending the analysis to other membersof the family, yeast Aac2p and mouse uncoupling protein 2, suggestsa degree of conservation. Simulation of large patches of a model mitochondrialmembrane containing multiple copies of the translocase shows thatCL interactions persist in the presence of protein–proteininteractions and suggests CL may mediate interactions between translocases.This study provides a key example of how computational microscopymay be used to shed light on regulatory lipid–protein interactions.

No MeSH data available.


Atomistic simulationof bovine ANT1. (A) The left panel is a viewfrom the matrix side, with the phosphate particles of the bound CLmolecules shown as spheres. For each phosphate particle, 1000 evenlydistributed snapshots are shown over the 0.5 μs simulation,colored according to simulation time. This may be compared to thephosphate distribution (right) for a noninteracting bulk CL. (B) Finalsnapshot of site I showing the arrangement of conserved motifs (gray),N-terminal ends of the helices (green and cyan), and CL (magenta).
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fig4: Atomistic simulationof bovine ANT1. (A) The left panel is a viewfrom the matrix side, with the phosphate particles of the bound CLmolecules shown as spheres. For each phosphate particle, 1000 evenlydistributed snapshots are shown over the 0.5 μs simulation,colored according to simulation time. This may be compared to thephosphate distribution (right) for a noninteracting bulk CL. (B) Finalsnapshot of site I showing the arrangement of conserved motifs (gray),N-terminal ends of the helices (green and cyan), and CL (magenta).

Mentions: To further assess interactionswithin the identified binding sites,a CG snapshot of ANT1 was converted42 toatomistic detail and simulated for 0.5 μs. Sites I and II ofthe chosen snapshot were occupied by CL molecules, while site IIIwas unoccupied, thus allowing us both to test the stability of boundCL molecules and to explore possible atomistic binding events withinthe same extended simulation. Calculation of the Cα root-mean-squaredeviation (RMSD) during the simulation revealed a stable value of∼0.3 nm for the core structural elements (i.e., helices) and∼0.4 nm for all residues, indicating that, despite the absenceof the carboxyatractyloside inhibitor, the apo-cytoplasm-facing structureis conformationally stable on the time scale simulated (Figure S6). The CL molecules within sites I andII remained bound (Figure 4), with the phosphate moieties contacting those conservedmotifs seen to form frequent contacts in the CG simulations. In contrast,the acyl chain tails exhibited more dynamic interactions and adopteda range of conformations (Figure S6,B).Of interest within site I, the side chain of R71 remained pointingdownward, consistent with its position within crystal structures (Figure 4B).9 No lipid binding events were observed at site III, despiteone CL being positioned in the vicinity of the site. This suggestslonger time scales may be required to capture the full process oflipid binding at atomistic resolution.


Lipid-Loving ANTs: Molecular Simulations of CardiolipinInteractions and the Organization of the Adenine Nucleotide Translocase in Model Mitochondrial Membranes
Atomistic simulationof bovine ANT1. (A) The left panel is a viewfrom the matrix side, with the phosphate particles of the bound CLmolecules shown as spheres. For each phosphate particle, 1000 evenlydistributed snapshots are shown over the 0.5 μs simulation,colored according to simulation time. This may be compared to thephosphate distribution (right) for a noninteracting bulk CL. (B) Finalsnapshot of site I showing the arrangement of conserved motifs (gray),N-terminal ends of the helices (green and cyan), and CL (magenta).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5120876&req=5

fig4: Atomistic simulationof bovine ANT1. (A) The left panel is a viewfrom the matrix side, with the phosphate particles of the bound CLmolecules shown as spheres. For each phosphate particle, 1000 evenlydistributed snapshots are shown over the 0.5 μs simulation,colored according to simulation time. This may be compared to thephosphate distribution (right) for a noninteracting bulk CL. (B) Finalsnapshot of site I showing the arrangement of conserved motifs (gray),N-terminal ends of the helices (green and cyan), and CL (magenta).
Mentions: To further assess interactionswithin the identified binding sites,a CG snapshot of ANT1 was converted42 toatomistic detail and simulated for 0.5 μs. Sites I and II ofthe chosen snapshot were occupied by CL molecules, while site IIIwas unoccupied, thus allowing us both to test the stability of boundCL molecules and to explore possible atomistic binding events withinthe same extended simulation. Calculation of the Cα root-mean-squaredeviation (RMSD) during the simulation revealed a stable value of∼0.3 nm for the core structural elements (i.e., helices) and∼0.4 nm for all residues, indicating that, despite the absenceof the carboxyatractyloside inhibitor, the apo-cytoplasm-facing structureis conformationally stable on the time scale simulated (Figure S6). The CL molecules within sites I andII remained bound (Figure 4), with the phosphate moieties contacting those conservedmotifs seen to form frequent contacts in the CG simulations. In contrast,the acyl chain tails exhibited more dynamic interactions and adopteda range of conformations (Figure S6,B).Of interest within site I, the side chain of R71 remained pointingdownward, consistent with its position within crystal structures (Figure 4B).9 No lipid binding events were observed at site III, despiteone CL being positioned in the vicinity of the site. This suggestslonger time scales may be required to capture the full process oflipid binding at atomistic resolution.

View Article: PubMed Central - PubMed

ABSTRACT

The exchange of ADPand ATP across the inner mitochondrial membraneis a fundamental cellular process. This exchange is facilitated bythe adenine nucleotide translocase, the structure and function ofwhich are critically dependent on the signature phospholipid of mitochondria,cardiolipin (CL). Here we employ multiscale molecular dynamics simulationsto investigate CL interactions within a membrane environment. Usingsimulations at both coarse-grained and atomistic resolutions, we identifythree CL binding sites on the translocase, in agreement with thoseseen in crystal structures and inferred from nuclear magnetic resonancemeasurements. Characterization of the free energy landscape for laterallipid interaction via potential of mean force calculations demonstratesthe strength of interaction compared to those of binding sites onother mitochondrial membrane proteins, as well as their selectivityfor CL over other phospholipids. Extending the analysis to other membersof the family, yeast Aac2p and mouse uncoupling protein 2, suggestsa degree of conservation. Simulation of large patches of a model mitochondrialmembrane containing multiple copies of the translocase shows thatCL interactions persist in the presence of protein–proteininteractions and suggests CL may mediate interactions between translocases.This study provides a key example of how computational microscopymay be used to shed light on regulatory lipid–protein interactions.

No MeSH data available.