A re-evaluation of energy-independent calcium-ion binding by rat liver mitochondria.

The impermeability of the mitochondrial inner membrane to the chelator ethanedioxybis(ethylamine)tetra-acetic acid permits discrimination between Ca(2+) which has been transported to the internal (matrix) phase and Ca(2+) which binds to the external surfaces of the mitochondrion. With this technique, it is shown that ;energy-independent high-affinity' binding is a measure of carrier-mediated active Ca(2+) transport in respiration-inhibited mitochondria; the carrier also transports Ca(2+) to the internal phase after treatment with carbonyl cyanide m-chlorophenylhydrazone, but in this case the active-transport component is inhibited. The Ca(2+)-binding sites associated with the external membrane surfaces are similar in concentration and affinity for both inhibited and uncoupled mitochondria; it was not possible to measure external Ca(2+) binding which could be identified as carrier specific. The results are discussed in relation to the mechanism of mitochondrial Ca(2+) transport, and to previous studies of energy-independent Ca(2+) binding.