ATP transport through a single mitochondrial channel, VDAC, studied by current fluctuation analysis.
The "molecular Coulter counter" concept has been used to study transport of ATP molecules through the nanometer-scale aqueous pore of the voltage-dependent mitochondrial ion channel, VDAC. We examine the ATP-induced current fluctuations and the change in average current through a single fully open channel reconstituted into a planar lipid bilayer. At high salt concentration (1 M NaCl), the addition of ATP reduces both solution conductivity and channel conductance, but the effect on the channel is several times stronger and shows saturation behavior even at 50 mM ATP concentration. These results and simple steric considerations indicate pronounced attraction of ATP molecules to VDAC's aqueous pore and permit us to evaluate the effect of a single ATP molecule on channel conductance. ATP addition also generates an excess noise in the ionic current through the channel. Analysis of this excess noise shows that its spectrum is flat in the accessible frequency interval up to several kilohertz. ATP exchange between the pore and the bulk is fast enough not to display any dispersion at these frequencies. By relating the low-frequency spectral density of the noise to the equilibrium diffusion of ATP molecules in the aqueous pore, we calculate a diffusion coefficient D = (1.6-3.3)10(-11) m2/s. This is one order of magnitude smaller than the ATP diffusion coefficient in the bulk, but it agrees with recent results on ATP flux measurements in multichannel membranes using the luciferin/luciferase method.
Rostovtseva
T K
TK
Laboratory of Physical and Structural Biology, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892, USA.
Bezrukov
S M
SM
D000255:Adenosine Triphosphate / Q000378:metabolism* / Q000494:pharmacology; D004058:Diffusion; D007425:Intracellular Membranes / Q000502:physiology*; D007700:Kinetics; D008051:Lipid Bilayers; D008564:Membrane Potentials / Q000187:drug effects; D008565:Membrane Proteins / Q000737:chemistry / Q000187:drug effects / Q000502:physiology*; D008928:Mitochondria / Q000502:physiology*; D008956:Models, Chemical; D008958:Models, Molecular; D009492:Neurospora crassa / Q000502:physiology; D018272:Porins; D017433:Protein Structure, Secondary; D012462:Saline Solution, Hypertonic / Q000494:pharmacology; D050994:Voltage-Dependent Anion Channels
D008051:Lipid Bilayers; D008565:Membrane Proteins; D018272:Porins; D012462:Saline Solution, Hypertonic; D050994:Voltage-Dependent Anion Channels; D000255:Adenosine Triphosphate