Voltage-dependent anion selective channel isoform1 maintains the permeability of the outer mitochondrial membrane. Its voltage-gating properties are relevant in bioenergetic metabolism and apoptosis. The N-terminal domain is suspected to be involved in voltage-gating, due to its peculiar localization. However this issue is still controversial. In this work we exchanged or deleted the beta-strands that take contact with the N-terminal domain. The exchange of the whole hVDAC1 beta-barrel with the homologous hVDAC3 beta-barrel produces a chimeric protein that, in reconstituted systems, loses completely voltage-dependence. hVDAC3 beta-barrel has most residues in common with hVDAC1, including V143 and L150 considered anchor points for the N-terminus. hVDAC1 mutants completely lacking either the beta-strand 9 or both beta-strands 9 and 10 were expressed, refolded and reconstituted in artificial bilayers. The mutants formed smaller pores. Molecular dynamics simulations of the mutant structure supported its ability to form smaller pores. The mutant lacking both beta-strands 9 and 10 showed a new voltage-dependence feature resulting in a fully asymmetric behavior. These data indicate that a network of beta-strands in the pore-walls, and not single residues, are required for voltage-gating in addition to the N-terminus. (C) 2013 Elsevier B.V. All rights reserved.
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