Flexibility of the N-Terminal mVDAC1 Segment Controls the Channel’s Gating Behavior

Abstract

Since the solution of the molecular structures of members of the voltage dependent anion channels (VDACs), the N-terminal α-helix has been the main focus of attention, since its strategic location, in combination with its putative conformational flexibility, could define or control the channel’s gating characteristics. Through engineering of two double-cysteine mVDAC1 variants we achieved fixing of the N-terminal segment at the bottom and midpoint of the pore. Whilst cross-linking at the midpoint resulted in the channel remaining constitutively open, cross-linking at the base resulted in an “asymmetric” gating behavior, with closure only at one electric field´s orientation depending on the channel’s orientation in the lipid bilayer. Additionally, and while the native channel adopts several well-defined closed states (S1 and S2), the cross-linked variants showed upon closure a clear preference for the S2 state. With native-channel characteristics restored following reduction of the cysteines, it is evident that the conformational flexibility of the N-terminal segment plays indeed a major part in the control of the channel’s gating behavior.

Voltage-dependent structural changes of the membrane-bound anion channel hVDAC1 probed by SEIRA and electrochemical impedance spectroscopy

“…18 Accordingly, the a-helix may not only passively support the global structural changes of the pore. Instead, both the b-barrel (model 1) or the bilayer (model 2) and the a-helix may respond to the electric field and contribute to the opening and closing events of the pore in a synergetic manner.…”

“…[13][14][15][16] These theories involved, in particular, movements of the helix into the centre of the pore or realignments of the b-sheet structure to change the pore size or shape, 13,14 or the electrical properties of the lumen of VDAC. 17,18 Recent studies focused on the role of the a-helix by fixing it covalently to the barrel wall. 18,19 Mertins et al 18 demonstrated that fixing the helix at two different positions either impaired voltage-induced closure of the channel or produced an asymmetric gating that depended on the direction of the electric field.…”

Nucleotide Interactions of the Human Voltage-dependent Anion Channel

“…Electron microscopy and bilayer measurements suggest moderate to large structural changes during channel closure (5,9,20). Models for gating mechanisms include rearrangements of the N-terminal ␣-helix, barrel deformations, a combination of helix and barrel motions to varying extent, and translocation of a membrane-embedded voltage sensor out of the membrane (4,(21)(22)(23)(24)(25)(26)(27). Although NADH enhances the channel voltage dependence and reduces nucleotide permeability of the outer mitochondrial membrane, ATP itself has no effect on VDAC gating (16,28).…”

Modulation of the mitochondrial voltage dependent anion channel (VDAC) by curcumin

“…This mutant showed lesser sensitivity to curcumin. N-terminal α helix of VDAC has been implicated in gating [4,8,9]. Although not universally accepted, it is generally believed that transition from closed to open state is accompanied by the conformational movement of N-terminus.…”

“…Open state is anion selective while closed state prefers cations [4,8]. Although there are controversies, large number of evidences support that the N-terminus participates in the gating of VDAC [8][9][10][11][12]. Flexibility and strategic location of the α helix within the pore prompted the hypothesis that channel opening is accompanied by the displacement of the helix from its central position to the inner wall [13].…”