Abstract
Molecular dynamics (MD) simulations of uncoupling proteins (UCP), a class of transmembrane proteins relevant for proton transport across inner mitochondrial membranes, represent a complicated task due to the lack of available structural data. In this work, we use a combination of homology modelling and subsequent microsecond molecular dynamics simulations of UCP2 in the DOPC phospholipid bilayer, starting from the structure of the mitochondrial ATP/ADP carrier (ANT) as a template. We show that this protocol leads to a structure that is impermeable to water, in contrast to MD simulations of UCP2 structures based on the experimental NMR structure. We also show that ATP binding in the UCP2 cavity is tight in the homology modelled structure of UCP2 in agreement with experimental observations. Finally, we corroborate our results with conductance measurements in model membranes, which further suggest that the UCP2 structure modeled from ANT protein possesses additional key functional elements, such as a fatty acid-binding site at the R60 region of the protein, directly related to the proton transport mechanism across inner mitochondrial membranes.
Highlights
Uncoupling protein 2 (UCP2) belongs to the mitochondrial SLC25 superfamily of anion transporters
We show that this protocol leads to a structure that is impermeable to water, in contrast to Molecular dynamics (MD) simulations of UCP2 structures based on the experimental NMR structure
We corroborate our results with conductance measurements in model membranes, which further suggest that the UCP2 structure modeled from ANT protein possesses additional key functional elements, such as a fatty acid-binding site at the R60 region of the protein, directly related to the proton transport mechanism across inner mitochondrial membranes
Summary
Uncoupling protein 2 (UCP2) belongs to the mitochondrial SLC25 superfamily of anion transporters. Motivated by the lack of relevant MD simulations for the monomeric structures which would help to decipher the function of UCP2 protein in membranes, we turned to homology modeling using the structure of mitochondrial ADP/ATP carrier We should mention that in the case of UCP2h and ANT structures, the distances between pairs of negatively charged residues at the matrix side (i.e., EGmotif), which were highly conserved across mitochondrial ADP/ATP carriers [47], kept three-fold pseudosymmetry in contrast to the UCP2NMR structure where this motif was not conserved, and distances between the negatively charged residues were larger (Figure S3) In this way, we further showed that the UCP2NMR structure was unstable and functionally irrelevant when embedded in phospholipid bilayers, which were structurally significantly different compared to the alkyl phosphocholine environment serving as an extracting agent [20,22,33]. A top-down view on the matrix exposed side of selected protein snapshots of UCP2NMR and UCP2h structures is shown on the right
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