Biogenesis of Cytochrome c in Neurospora crassa

# 1. Precipitating antibodies specific for apocytochrome c and
holocytochrome c, respectively, were employed to study synthesis
and intracellular transport of cytochrome c in Neurospora in vitro.
# 2. Apocytochrome c as well as holocytochrome c were found to be
synthesized in a cell-free homogenate. A precursor product
relationship between the two components is suggested by kinetic
experiments. # 3. Apocytochrome c synthesized in vitro was found in
the post-ribosomal fraction and not in the mitochondrial fraction,
whereas holocytochrome c synthesized in vitro was mainly detected
in the mitochondrial fraction. A precursor product relationship
between postribosomal apocytochrome c and mitochondrial
holocytochrome c is indicated by the labelling data. In the
microsomal fraction both apocytochrome c and holocytochrome c were
found in low amounts. Their labelling kinetics do not suggest a
precursor role of microsomal apocytochrome c or holocytochrome c. #
4. Formation of holocytochrome c from apocytochrome c was observed
when postribosomal supernatant containing apocytochrome c
synthesized in vitro was incubated with isolated mitochondria, but
not when incubated in the absence of mitochondria. The cytochrome c
formed under these conditions was detected in the mitochondria. #
5. Conversion of labelled apocytochrome c synthesized in vitro to
holocytochrome c during incubation of a postribosomal supernatant
with isolated mitochondria was inhibited when excess isolated
apocytochrome c, but not when holocytochrome c was added. # 6. The
data presented are interpreted to show that apocytochrome c is
synthesized on cytoplasmic ribosomes and released into the
supernatant. It is suggested that apocytochrome c migrates to the
inner mitochondrial membrane, where the heme group is covalently
linked to the apoprotein. The hypothesis is put forward that the
concomitant change in conformation leads to trapping of
holocytochrome c in the membrane. The probles of permeability of
the outer mitochondrial membrane to apocytochrome c and the site
and nature of the reaction by which the heme group is linked to the
apoprotein are discussed.