Circular dichroism studies of the mitochondrial channel, VDAC, from Neurospora crassa.
The protein that forms the voltage-gated channel VDAC (or mitochondrial porin) has been purified from Neurospora crassa. At room temperature and pH 7, the circular dichoism (CD) spectrum of VDAC suspended in octyl beta-glucoside is similar to those of bacterial porins, consistent with a high beta-sheet content. When VDAC is reconstituted into phospholipid liposomes at pH 7, a similar CD spectrum is obtained and the liposomes are rendered permeable to sucrose. Heating VDAC in octyl beta-glucoside or in liposomes results in thermal denaturation. The CD spectrum irreversibly changes to one consistent with total loss of beta-sheet content, and VDAC-containing liposomes irreversibly lose sucrose permeability. When VDAC is suspended at room temperature in octyl beta-glucoside at pH < 5 or in sodium dodecyl sulfate at pH 7, its CD spectrum is consistent with partial loss of beta-sheet content. The sucrose permeability of VDAC-containing liposomes is decreased at low pH and restored at pH 7. Similarly, the pH-dependent changes in the CD spectrum of VDAC suspended in octyl beta-glucoside also are reversible. These results suggest that VDAC undergoes a reversible conformational change at low pH involving reduced beta-sheet content and loss of pore-forming activity.
Shao
L
L
Division of Molecular Medicine, Wadsworth Center, Albany, New York, USA.
Kinnally
K W
KW
Mannella
C A
CA
D000596:Amino Acids / Q000032:analysis; D015153:Blotting, Western; D002942:Circular Dichroism; D004594:Electrophysiology; D006863:Hydrogen-Ion Concentration; D015640:Ion Channel Gating; D007473:Ion Channels / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D008081:Liposomes; D008565:Membrane Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000502:physiology*; D008928:Mitochondria / Q000502:physiology*; D009492:Neurospora crassa / Q000502:physiology*; D018272:Porins; D011487:Protein Conformation; D013056:Spectrophotometry, Ultraviolet; D050994:Voltage-Dependent Anion Channels
D000596:Amino Acids; D007473:Ion Channels; D008081:Liposomes; D008565:Membrane Proteins; D018272:Porins; D050994:Voltage-Dependent Anion Channels