Voltage-dependent anion channel transports calcium ions through biomimetic membranes.
The mitochondrial outer membrane channel (VDAC), a central player in mitochondria and cell death, was reconstituted in polymer-supported phospholipid bilayers. Highly purified VDAC was first reconstituted in vesicles; channel properties and NADH-ferricyanide reductase activity were ascertained before deposition onto solid substrates. 1-Palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine/1,2-distearoyl-sn-glycero-3-phosphoethanolamine-poly(ethylene glycol)-N-hydroxysuccinimide mixed vesicles containing VDAC were linked onto amine-grafted surfaces (glass and gold) and disrupted to form a VDAC-containing polymer-tethered planar bilayer. Surface plasmon spectroscopy, fluorescence microscopy, and atomic force microscopy measurements ascertained the membrane thickness, fluidity, and continuity. VDAC reconstituted in bilayers efficiently transported calcium ions and was modulable by two channel blockers, 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid and l-glutamate. The novel setup may allow the study of the assembly of a polyprotein complex centered on VDAC and its role in mitochondrial biology, calcium fluxes, and apoptosis.
Deniaud
Aurélien
A
CNRS UMR 8159, Laboratoire de Génétique et Biologie Cellulaire, Université de Versailles/St Quentin, 45 Avenue des Etats-Unis, 78035 Versailles, France.
Rossi
Claire
C
Berquand
Alexandre
A
Homand
Johanne
J
Campagna
Sylvie
S
Knoll
Wolfgang
W
Brenner
Catherine
C
Chopineau
Joël
J
D000818:Animals; D017209:Apoptosis; D002118:Calcium / Q000737:chemistry*; D002121:Calcium Channel Blockers / Q000737:chemistry; D006801:Humans; D017136:Ion Transport; D008567:Membranes, Artificial; D046912:Multiprotein Complexes / Q000737:chemistry*; D009247:NADH, NADPH Oxidoreductases / Q000737:chemistry; D020349:Surface Plasmon Resonance; D050994:Voltage-Dependent Anion Channels / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000302:isolation & purification
D002121:Calcium Channel Blockers; D008567:Membranes, Artificial; D046912:Multiprotein Complexes; D050994:Voltage-Dependent Anion Channels; D009247:NADH, NADPH Oxidoreductases; C014227:ferricyanide reductase; D002118:Calcium