Human mitochondrial VDAC functionality governs scaffold stability

Abstract

ATP transport across the mitochondrial outer membrane occurs through voltage-dependent anion channels (VDACs). The three human VDAC isoforms share a similar primary sequence but differ in their interactome, and thereby, show antagonistic physiological functions. The anti-apoptotic and regulatory role of human VDAC2 (hV2) isoform must therefore originate from subtle variations in its primary sequence. However, the molecular details of how stability and function are regulated in hV2, and its impact on protein-protein interaction, are unknown. Here, we show that the 19-stranded hV2 barrel has evolutionarily retained an energetically suboptimal sequence for superior channel gating and voltage sensing. We find that residues in the N- and C-terminal zones, comprising strands β2-β8 and β17-β18, lower the scaffold stability by 1.0-3.0 kcal/mol. Furthermore, strands β5-β10 and β17 possess sequences that are intrinsically prone to association and aggregation. However, hV2 trades-off stability and aggregation for function. Residues in the N-terminal zone are important for voltage-dependent channel gating characteristics that are essential for metabolite transport. Our results demonstrate how energetic contribution of specific residues in hV2 links inversely to its functional importance. We conclude that the evolutionary selection of hV2 sequence for channel function and regulation is at the expense of innate barrel stability. To our knowledge, this is the first study to establish stability-function trade-off in a human β-barrel membrane protein. We propose that the antagonistic behavior of the three VDAC isoforms resides in subtle differences in the molecular elements that we have identified. These molecular elements decide thermodynamic stability, function, and the association network of the VDAC channels in the mitochondrial outer membrane.