Molecular Design of the Voltage-Dependent, Anion-Selective Channel in the Mitochondrial Outer Membrane

Guo, Xiao; Smith, PR; Cognon, B.; D'Arcangelis, D.; Dolginova, Elena A.; Mannella, Carmen A.

doi:10.1006/jsbi.1995.1004

Cited by 92 publications

(94 citation statements)

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“…During voltage-induced gating, the N-terminal region either stays inside the pore or moves in a concerted fashion with the rest of the voltage sensor rather than exerting large independent structural rearrangements to achieve voltage-dependent gating. It should be noted that the data presented here still permit the translocation of the N terminus in and out of the lumen upon gating, as suggested previously (16,20,32,46), as long as this movement is coordinated with the voltage sensor.If voltage gating does not involve rearrangements of the N-terminal segment, then how is voltage gating achieved? There is still no clear structural understanding of VDAC dynamics, but a new mechanism is emerging; the barrel wall becomes elongated, and the pore narrows around the intact N-terminal segment to exclude nucleotides.…”

supporting

confidence: 78%

“…Based on these data, they developed a model for VDAC voltage gating where the N-terminal segment is an essential component of the mobile voltage sensor domain, which slides in and out of the channel lumen in response to the applied voltage (1,14,15). Several additional studies have reported alternative models in which the N-terminal segment is exposed to the cytoplasm (7,17,18) or resides on the membrane surface (19,20). Although the proposed mechanism of voltage gating differs among investigators, most models involve large conformational changes of the N-terminal segment.…”

mentioning

confidence: 99%

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Affixing N-terminal α-Helix to the Wall of the Voltage-dependent Anion Channel Does Not Prevent Its Voltage Gating

Teijido

Ujwal

Hillerdal

et al. 2012

Journal of Biological Chemistry

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Background: There is ongoing controversy concerning the location and mobility of the N-terminal ␣-helix in VDAC1 during voltage gating. Results: mVDAC1 with the N-terminal ␣-helix cross-linked to ␤-strand 11 forms typical voltage-gated channels. Conclusion:The N-terminal domain of VDAC1 does not move independently during voltage gating. Significance: This study dramatically alters the current view of voltage gating dynamic in VDAC1.The voltage-dependent anion channel (VDAC) governs the free exchange of ions and metabolites between the mitochondria and the rest of the cell. The three-dimensional structure of VDAC1 reveals a channel formed by 19 ␤-strands and an N-terminal ␣-helix located near the midpoint of the pore. The position of this ␣-helix causes a narrowing of the cavity, but ample space for metabolite passage remains. The participation of the N-terminus of VDAC1 in the voltage-gating process has been well established, but the molecular mechanism continues to be debated; however, the majority of models entail large conformational changes of this N-terminal segment. Here we report that the pore-lining N-terminal ␣-helix does not undergo independent structural rearrangements during channel gating. We engineered a double Cys mutant in murine VDAC1 that cross-links the ␣-helix to the wall of the ␤-barrel pore and reconstituted the modified protein into planar lipid bilayers. The modified murine VDAC1 exhibited typical voltage gating. These results suggest that the N-terminal ␣-helix is located inside the pore of VDAC in the open state and remains associated with ␤-strand 11 of the pore wall during voltage gating.The voltage-dependent anion channel (VDAC) 4 serves as the primary conduit between the mitochondria and the rest of the cell, facilitating free exchange of ions and metabolites across the outer mitochondrial membrane. In addition to its metabolic and energetic functions, VDAC has a more complex role, serving as a receptor for molecules and proteins that modulate the organelle's permeability and thereby its function (1-3). This multifunctional channel has been implicated in the metabolic stresses of cancer, cardiovascular disease, and mitochondrialdependent cell death (4 -8); thus, understanding the structure and function of VDAC constitutes a critical objective for basic as well as medical research.Single channel conductance experiments on VDAC1 at low membrane potential (Ͻ30 mV) reveal a high conductance (4.1 Ϯ 0.1 nanosiemens in 1 M KCl) indicative of a large pore, usually referred to as the "open" state of the channel (1). This conformer facilitates the passage of 10 6 ATP molecules (9) and displays a preference for monovalent anions over cations with the anion-to-cation permeability ratio of 2:1 in high salt (10) and 4:1 in physiological salt concentration (11). As voltage is increased (Ͼ30 mV) in either a positive or a negative direction, the channel switches into the lower conducting states (around 2 nanosiemens in 1 M KCl), termed as the "closed" state(s). These states are cation-selective wi...

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supporting

confidence: 78%

mentioning

confidence: 99%

Affixing N-terminal α-Helix to the Wall of the Voltage-dependent Anion Channel Does Not Prevent Its Voltage Gating

Teijido

Ujwal

Hillerdal

et al. 2012

Journal of Biological Chemistry

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“…7 would suggest that these sites should be on opposite sides of the membrane. Previous studies have demonstrated that this N-terminal ␣ helix is a mobile domain that may partition preferentially to one side of the membrane (21). The model shown in Fig.…”

Section: Discussionmentioning

confidence: 99%

The Topology of VDAC as Probed by Biotin Modification

Song

Midson

Blachly-Dyson

et al. 1998

Journal of Biological Chemistry

109

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The outer membrane of mitochondria contains channels called VDAC (mitochondrial porin), which are formed by a single 30-kDa protein. Cysteine residues introduced by site-directed mutagenesis at sites throughout Neurospora crassa VDAC (naturally devoid of cysteine) were specifically biotinylated prior to reconstitution into planar phospholipid membranes. From previous studies, binding of streptavidin to single biotinylated sites results in one of two effects: reduced single-channel conductance without blockage of voltage gating (type 1) or locking of the channels in a closed conformation (type 2). All sites react with streptavidin only from one side of the membrane. Here, we extend this approach to VDAC molecules containing two cysteines and determine the location of each biotinylated residue with respect to the other within the membrane. When a combination of a type 1 and a type 2 site was used, each site could be observed to react with streptavidin. Two sets of sites located on opposite surfaces of the membrane were identified, thereby establishing the transmembrane topology of VDAC. A revised folding pattern for VDAC, consisting of 1 ␣ helix and 13 ␤ strands, is proposed by combining these results with previously obtained information on which sites are lining the aqueous pore.VDAC channels are found in the mitochondrial outer membrane of cells from all eukaryotic kingdoms (1). They are the major pathways by which metabolites, including ATP, pass through this outer membrane (2, 3) and, therefore, are likely to play important roles in the regulation of mitochondrial functions.Each VDAC channel consisting of a single 30-kDa polypeptide (4, 5) forms an aqueous pore ϳ3 nm in diameter (6, 7). Detailed information on the voltage-gating properties of VDAC and the associated conformational rearrangements have been obtained from studies of channels reconstituted into planar phospholipid membranes. These studies have demonstrated that, in response to transmembrane potentials of 30 mV or above, part of the channel wall, which serves as the voltage sensor, moves out of the membrane resulting in channel closure (8 -10). These closed channels are essentially impermeable to ATP and have a reduced permeability to organic anions (2, 11). However, they are still permeable to small non-electrolytes (up to 1.8 nm in diameter) and to small monovalent cations (12). VDAC can close in response to transmembrane potentials of both polarities, due to the presence of two different gating processes.The transmembrane topology of VDAC molecules has yet to be firmly established. Theoretical considerations predicted a "␤-barrel" structure (13,14). "Sided" ␤ strands that would be appropriate to separate an apolar from a polar environment, were tested by the use of site-directed mutations (15). If engineered charge changes in proposed transmembrane segments affected channel selectivity, the associated protein segment was proposed to be a transmembrane strand. A lack of effect resulted in assignment of the segment to the membrane surface. In thes...

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“…1B) (35). Moreover, antibodies raised against the N terminus of scVDAC were able to bind to these membranes only when mitochondrial envelopes were disrupted (36). The combined data suggest that the C terminus of VDAC extends into the IMS, resembling the orientation found in all bacterial OMPs (24).…”

Section: Dimerization Of Vdac Via a Small Hydrophobic Interfacementioning

confidence: 94%

Structure of the human voltage-dependent anion channel

Bayrhuber

Meins

Habeck

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

508

626

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The voltage-dependent anion channel (VDAC), also known as mitochondrial porin, is the most abundant protein in the mitochondrial outer membrane (MOM). VDAC is the channel known to guide the metabolic flux across the MOM and plays a key role in mitochondrially induced apoptosis. Here, we present the 3D structure of human VDAC1, which was solved conjointly by NMR spectroscopy and x-ray crystallography. Human VDAC1 (hVDAC1) adopts a ␤-barrel architecture composed of 19 ␤-strands with an ␣-helix located horizontally midway within the pore. Bioinformatic analysis indicates that this channel architecture is common to all VDAC proteins and is adopted by the general import pore TOM40 of mammals, which is also located in the MOM.T he outer membrane of mitochondria (MOM) contains three integral membrane protein families, two of which form channels as part of larger protein complexes (for review, see ref. 1). These two MOM complexes, the general import pore TOM and the SAM insertase, allow for the entire translocation and insertion of nearly all newly synthesized proteins destined to the mitochondrial organelle (2, 3). The third protein family of typically high abundance (Ϸ10,000 copies per mitochondrion) is termed voltage-dependent anion channels (VDACs), because of the voltage sensitivity of its open probability (4, 5). Together, this small number of protein families is sufficient for full communication between mitochondria with their cellular environment (1).The VDAC channel was initially described as being reminiscent of bacterial porins and primarily responsible for the exchange of chemical energy equivalents between the cytosol and the mitochondrion (4, 6). Indeed, a variety of structural features (like barrel geometry and dimension) known from the bacterial precursors are maintained (7,8). By contrast, a variety of functions have been ascribed to the VDAC isoforms among which the direct coupling of the mitochondrial matrix to the energy maintenance of the cytosol seems to be the most general function (9). The structure of VDAC is of interest because of a substantial body of evidence connecting VDAC to apoptosis. It is suggested that VDAC is a critical player in the release of apoptogenic factors from mitochondria of mammalian cells, and consequently several hypotheses describing the mechanism of mitochondria-mediated apoptosis involving VDAC have been proposed (for review, see ref. 10). Results and DiscussionStructure Determination of hVDAC1: Combining NMR Spectroscopy and X-Ray Crystallography. In a parallel structural biology approach, we set out to characterize the structure of hVDAC1, the major isoform of this channel in mammalian tissues, by a combination of NMR spectroscopy and x-ray crystallography. The idea behind this project was to gain complementary structural information to have a solid basis for future studies, e.g., analysis of protein heterocomplex formation by NMR and crystal structures as a basis for drug target design. Only information derived from both methods and the application of an iterative s...

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Molecular Design of the Voltage-Dependent, Anion-Selective Channel in the Mitochondrial Outer Membrane

Cited by 92 publications

References 0 publications

Affixing N-terminal α-Helix to the Wall of the Voltage-dependent Anion Channel Does Not Prevent Its Voltage Gating

Affixing N-terminal α-Helix to the Wall of the Voltage-dependent Anion Channel Does Not Prevent Its Voltage Gating

The Topology of VDAC as Probed by Biotin Modification

Structure of the human voltage-dependent anion channel

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