Protein folding causes an arrest of preprotein translocation into mitochondria in vivo

With vital yeast cells, a hybrid protein consisting of the amino-
terminal third of the precursor to cytochrome b2 and of the entire
dihydrofolate reductase was arrested on the import pathway into
mitochondria. Accumulation of the protein in the mitochondrial
membranes was achieved by inducing a stable tertiary structure of
the dihydrofolate reductase domain. Thereby, three salient features
of mitochondrial protein uptake in vivo were demonstrated: its
posttranslational character; the requirement for unfolding of
precursors; and import through translocation contact sites. The
permanent occupation of translocation sites by the fusion protein
inhibited the import of other precursors; it did, however, not lead
to leakage of mitochondrial ions, implying the existence of a
channel that is sealed around the membrane spanning polypeptide
segment.