The topology of VDAC as probed by biotin modification.

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).

The voltage-gating process of the voltage-dependent anion channel is sensitive to ion flow.

The voltage-dependent anion channel (VDAC) is a voltage-gated channel from the mitochondrial outer membrane. It has two gating processes: one at positive potentials and the other at negative potentials. The energetics of VDAC gating are quite different when measured in the presence or absence of an ion gradient.

The sensor regions of VDAC are translocated from within the membrane to the surface during the gating processes.

The motion of the sensor regions in a mitochondrial voltage-gated channel called VDAC were probed by attaching biotin at specific locations and determining its ability to bind to added streptavidin. Site-directed mutagenesis was used to introduce single cysteine residues into Neurospora crassa VDAC (naturally lacks cysteine). These were chemically biotinylated and reconstituted into planar phospholipid membranes.

Successful recovery of the normal electrophysiological properties of PorB (class 3) porin from Neisseria meningitidis after expression in Escherichia coli and renaturation.

Neisseria meningitidis PorB class 3 porins obtained either from native membranes (wild-type) or recovered from inclusion bodies following expression in Escherichia coli (recombinant), have been reconstituted into solvent-free planar phospholipid membranes. The wild-type and recombinant porins exhibited the same single-trimer conductance (1-1.3 nS in 200 mM NaCl), tri-level closure pattern, characteristic of functional channel trimers, and pattern of insertion into planar membranes.

The role of yeast VDAC genes on the permeability of the mitochondrial outer membrane.

In addition to the POR1 gene, which encodes the well-characterized voltage dependent anion-selective channel (YVDAC1) of the mitochondrial outer membrane, the yeast Saccharomyces cerevisiae contains a second gene (POR2) encoding a protein (YVDAC2) with 50% sequence identity to YVDAC1. Mitochondria isolated from yeast cells deleted for the POR1 gene (delta por1) had a profoundly reduced outer membrane permeability as measured by the ability of an intermembrane space dehydrogenase to oxidize exogenously added NADH. Mitochondria missing either YVDAC1 or both YVDAC1 and YVDAC2 showed a 2-fold increase in the rate of NADH oxidation when the outer membrane was deliberately damaged.

Multicopy suppressors of phenotypes resulting from the absence of yeast VDAC encode a VDAC-like protein.

The permeability of the outer mitochondrial membrane to most metabolites is believed to be based in an outer membrane, channel-forming protein known as VDAC (voltage-dependent anion channel). Although multiple isoforms of VDAC have been identified in multicellular organisms, the yeast Saccharomyces cerevisiae has been thought to contain a single VDAC gene, designated POR1. However, cells missing the POR1 gene (delta por1) were able to grow on yeast media containing a nonfermentable carbon source (glycerol) but not on such media at elevated temperature (37 degrees C).

Isolation and characterization of the mitochondrial channel, VDAC, from the insect Heliothis virescens.

A 31 kDa voltage-dependent anion-selective channel (VDAC) protein was purified from the insect Heliothis virescens (tobacco budworm, denoted TBW) using an alkali extraction and filtration procedure and was characterized by SDS-PAGE, amino acid sequencing, biophysical properties and immunocytochemistry. The N-terminal sequence has highest identity with VDACs from mammals (50-66%) followed by plants (34-41%) and lower eukaryotes (30-34%). Reconstitution in planar phospholipid membranes yielded properties typical of VDACs from other organisms including a single-channel conductance of 4.

Regulation of metabolite flux through voltage-gating of VDAC channels.

The mitochondrial outer membrane channel, VDAC, is thought to serve as the major permeability pathway for metabolite flux between the cytoplasm and mitochondria. The permeability of VDAC to citrate, succinate, and phosphate was studied in channels reconstituted into planar phospholipid membranes. All ions showed large changes in permeability depending on whether the channel was in the open or in the low conductance, "closed" state, with the closed state always more cation selective.

Autodirected insertion: preinserted VDAC channels greatly shorten the delay to the insertion of new channels.

VDAC, a mitochondrial outer membrane channel, has the ability to catalyze and direct the insertion of other VDAC channels into planar phospholipid membranes. The spontaneous rate of insertion of detergent-solubilized VDAC channels into phospholipid membranes is estimated to be 1.5 x 10(-5) channels min-1 micron-2.

VDAC channels mediate and gate the flow of ATP: implications for the regulation of mitochondrial function.

The mitochondrial channel, VDAC, forms large (3 nm in diameter) aqueous pores through membranes. We measured ATP flow (using the luciferin/luciferase method) through these channels after reconstitution into planar phospholipid membranes. In the open state of VDAC, as many as 2 x 10(6) ATP molecules can flow through one channel per second.

ATP flux is controlled by a voltage-gated channel from the mitochondrial outer membrane.

A voltage-gated channel, called VDAC (mitochondrial porin) is known to be responsible for most of the metabolite flux across the mitochondrial outer membrane. Here, direct measurements of ATP flux through VDAC channels reconstituted into planar phospholipid membranes establish that VDAC is sufficient to provide passage for ATP efflux from mitochondria. Further, the gating of the channel can shut down ATP flux completely while, simultaneously, allowing the flow of small ions.

Polychlorobiphenyls in Antarctica.

The presence and the distribution of polychlorobiphenyls (PCBs) in Antarctic environmental components and the effect of the seasonal formation/melting process of pack ice on the pollution level of seawater were investigated. Seawater, marine, and lake sediment and soil samples were collected in a large area of the Ross Sea and Victoria Land during the 1988-1989, 1989-1990, 1990-1991, and 1991-1992 Italian expeditions. The results obtained highlighted a low and quite homogeneous PCB contamination of the studied area.

Decomposition Products of s-Triazine Herbicides by Electron-Transfer in Acidic Aqueous Media.

The reduction of atrazine and terbutylazine was preceded by protonation equilibrium. Three protonation sites of the s-triazine molecule determined the structure of the final reduction product. Protonation was investigated by the change of UV-Vis spectra.

Photodegradation of Polychlorinated Dibenzo-p-dioxins in Liquid Samples by Near UV Light Irradiation.

Photodegradation of 1,2,3,4-tetrachlorodibenzo-p-dioxin (1,2,3,4-TCDD) in hexane solution was studied under controlled near-UV light exposure in the spectral region from 325 to 269 nm. GC-MS was used to detect the amount of unreacted dioxin and to characterize the relevant degradation products. Irradiation experiments carried out at a constant light energy (700 mJ) showed that the percentage of 1,2,3,4-TCDD left in the solution after irradiation changed from about 55 to 75%, with a minimum of 55% at 310 nm.

The role of pyridine dinucleotides in regulating the permeability of the mitochondrial outer membrane.

Both NADH and NADPH reduce the permeability of the mitochondrial outer membrane to ADP. This is specific for the outer membrane and uncorrelated with the respiratory control ratio. This could result in a 7-fold difference between the concentration of ADP in the intermembrane space and that in the external environment (at 5 microM ADP).

Self-catalyzed insertion of proteins into phospholipid membranes.

We present evidence that the mitochondrial channel, VDAC, when reconstituted into a phospholipid membrane, can catalyze the insertion of other VDAC channels. This property called "auto-directed insertion" was first proposed by Zizi et al. (1995) to explain observations on asymmetric VDAC channels.

The control of mitochondrial respiration in yeast: a possible role of the outer mitochondrial membrane.

Mitochondrial respiration in yeast (S. cerevisiae) is regulated by the level of glucose in the medium. Glucose is known to inhibit respiration by repressing key enzymes in the respiratory chain.

Individual leaflets of a membrane bilayer can independently regulate permeability.

Water rapidly crosses most membranes, but only slowly crosses apical membranes of barrier epithelia such as bladder and kidney collecting duct, a feature essential to barrier function. How apical membrane structure reduces permeabilities remains unclear. Cell plasma membranes contain two leaflets of distinct lipid composition; the role of this bilayer asymmetry in membrane permeability is unclear.

Indications of a common folding pattern for VDAC channels from all sources.

Previous research on the mitochondrial channel VDAC from the yeast S. cerevisiae had identified protein strands forming the wall of VDAC's aqueous pore. Here we report the results of analyzing the primary sequences of VDAC from various sources to see if the transmembrane folding pattern identified from this yeast is conserved for VDAC of different species.

Biliopancreatic diversion for obesity at eighteen years.

Background: Surgical attempts to treat obesity began because of the discouraging results of conservative medical treatment, which successfully achieved initial weight loss but failed to maintain it. Gastric restrictive procedures, currently the most popular surgical methods for obesity therapy, have proved to be effective in initiating weight loss, but some concerns regarding their long-term efficacy in weight maintenance have arisen.

Methods: Of a total of 1968 obese patients who underwent biliopancreatic diversion since 1976, the last consecutive 1217 underwent the "ad hoc stomach" type of diversion with a 200 cm alimentary limb, a 50 cm common limb, and a gastric volume varying between 200 and 500 ml.

Oriented channel insertion reveals the motion of a transmembrane beta strand during voltage gating of VDAC.

Yeast VDAC channels (isolated from the mitochondrial outer membrane) form large aqueous pores whose walls are believed to consist of 1 alpha helix and 12 beta strands. Each channel has two voltage-gating processes: one closes the channels at positive potentials, the other at negative. When VDAC is reconstituted into phospholipid (soybean) membranes, the two gating processes have virtually the same steepness of voltage dependence and the same midpoint voltage.

Beta-NADH decreases the permeability of the mitochondrial outer membrane to ADP by a factor of 6.

Mitochondria with intact outer membrane (99% intact based on cytochrome c impermeability) were isolated and used to measure the permeability of their outer membrane to ADP. beta-NADH reduced the permeability in a concentration-dependent manner (KD = 87 +/- 5 microM) by a factor of 6. alpha-NADH and beta-NAD+ cannot mimic the action of beta-NADH.

Well being in patients on CAPD and hemodialysis.

In the present multicenter study, 120 pts who had been treated by both hemodialysis (HD) and continuous ambulatory peritoneal dialysis (CAPD) for at least 6 months each, were invited to answer questions on 34 matters, to compare symptoms and their well-being while on the two treatments. Patients were invited to choose HD or CAPD and indicate the reasons for their choice. For 28 patients the first treatment was HD and for 92 CAPD.