Transport into mitochondria and intramitochondrial sorting of the Fe/S protein of ubiquinol-cytochrome c reductase

Protein transport into mitochondria

“…Membrane-permeable chelators, such as ortho-phenanthroline, inhibit the processing activity in intact mitochondria. Precursors can thus be imported into the mitochondrial matrix without the removal of their ami no-termina 1 presequences (11,22).…”

“…Major portions of the precursors, however, were still outside the outer membrane as they were accessible to proteases or to antibodies added to isolated mitochondria. Thus, the precursor proteins were reaching from the outside of the outer membrane into the matrix thereby spanning sites of close contact between both membranes (translocation contact sites) (3,8,(10)(11)(12). The translocation intermediates with bound antibodies were decorated with protein-A-gold particles.…”

“…The ability to block the processing peptidase is used as a tool to dissect the import pathway of these proteins into intermediate steps. Such experiments with the Rieske Fe/S-protein of the bc^-complex, a protein located at the outer surface of the inner membrane, had a surprising result (11,25). Following inhibition of the processing peptidase precursor of Fe/S-protein is accumulated as a soluble species in the mitochondrial matrix, as revealed by digitonin fractionation of mitochondria.…”

“…The precursors specifically interact with proteinaceous binding sites on the mitochondrial surface (5-9). Import into or across the inner membrane occurs at contact sites between both membranes (8,(10)(11)(12) and requires a membrane potential Afy across the inner membrane (13-16), Presequences are proteolytically removed by the processing peptidase located in the mitochondrial matrix (17)(18)(19)(20). The proteins are sorted to their final intramitochondrial location and are often assembled into multi-subunit complexes.…”

Contact sites between inner and outer membranes

“…tors of mitochondria [13,14]. Precursor proteins following this 'conservative sorting pathway' often possess a bipartite presequence the first part of which is cleaved off in the matrix and the second part directs the retranslation across the inner membrane.…”