Structural and functional analysis of human voltage-dependent anion channel isoforms (hVDACs): Combining in-vitro and in-silico approaches

VDACs are a small family of conserved proteins located in the outer mitochondrial membrane. They conduct ions, metabolites and small molecules, among which the energetic nucleotides ATP, ADP and NADH. Three different VDAC isoforms have been characterized in higher eukaryotes, encoded by three separate nuclear genes. VDAC1 is the most abundant isoform in most cells, being ten and hundred times more prevalent than VDAC2 and VDAC3, respectively. It is thus not surprising that VDAC1 is the isoform most extensively characterized. Functionally, VDAC1 is anion selective and exhibits a single-channel conductance of ~3.5-4.0 nS in 1 M KCl at an applied voltage between -20 mV and 10 mV. Raising the applied voltage results in the channel switching to the so-called “closed state”, with a lower conductance and a channel selectivity reversed to cations. In addition to the poreforming function, VDAC1 has been involved in various interactions and cross-talk with other cellular proteins like hexokinase, tubulin, the Ca2+ gate into mitochondria and the Bcl-2 family members that can impact on the activity of the pore itself and vice versa, testimony to the involvement of VDAC to crucial cell fates like in pathways leading to apoptosis, cancer and degeneration The aim of the PhD project was to perform a comparative study on the human VDAC isoforms focusing on both the whole channels and the individuals N-terminal domains. In this sense, both experimental and computational techniques have been used pointing out their complementarity and contribute to the completeness of the study. After a brief introduction, the methods used during the PhD will be presented. In the third chapter, the results together with the discussion will be described. Firstly, focusing on the structural characterization of the N-termini of the three isoforms. Secondly, the results and the discussion will concern the comparative study of the entire channels. Both of these characterization have been performed with either experimental and computational techniques. Finally, in the 4th chapter a brief conclusion and an outlook on a future perspective will be given.