The reversible Ca-induced permeabilization of rat liver mitochondria.
19 - 29.
Rat liver mitochondria became permeabilized to sucrose according to an apparent first-order process after accumulating 35 nmol of Ca/mg of protein in the presence of 2.5 mM-P(i), but not in its absence. A fraction (24-32%) of the internal space remains sucrose-inaccessible. The rate constant for permeabilization to sucrose decreases slightly when the pH is decreased from 7.5 to 6.5, whereas the rate of inner-membrane potential (Δψ) dissipation is markedly increased, which indicates that H permeation precedes sucrose permeation. Permeabilization does not release mitochondrial proteins. [C]Sucrose appears to enter permeabilized mitochondria instantaneously. Chelation of Ca with EGTA restores Δψ and entraps sucrose in the matrix space. With 20 mM-sucrose at the instant of resealing, about 21 nmol of sucrose/mg of protein becomes entrapped. The amount of sucrose entrapped is proportional to the degree of permeabilization. Entrapped sucrose is not removed by dilution of the mitochondrial suspension. Resealed mitochondria washed three times retain about 74% of the entrapped sucrose. In the presence of Ruthenium Red and Ca buffers permeabilized mitochondria reseal only partially with free [Ca] > 3 μM. [C]Sucrose enters partially resealed mitochondria continuously with time, despite maintenance of Δψ, in accordance with continued interconversion of permeable and impermeable forms. Kinetic analysis of [C]sucrose entry indicate two Ca-sensitive reactions in permeabilization. This conclusion is supported by the biphasic time courses of resealing and repolarization of permeabilized mitochondria and the acute dependence of the rapid repolarization on the free [Ca]. A hypothetical model of permeabilization and resealing is suggested and the potential of the procedure for matrix entrapment of substances is discussed.
|Title:||The reversible Ca-induced permeabilization of rat liver mitochondria|
|UCL classification:||UCL > School of Life and Medical Sciences > Faculty of Life Sciences > Biosciences (Division of)|
Archive Staff Only