Import of a mutant mitochondrial precursor fails to respond to stimulation by a cytosolic factor

Abstract

Import of the precursor to ornithine carbamyltransferase is stimulated by a partially‐purified, NEM‐sensitive soluble factor from rabbit reticulocyte lysate. A mutant in which the carboxy‐terminal 73 amino acids were deleted, had a sharply reduced response to this factor. The NEM‐sensitive, import‐stimulating factor interacts with the surface of mitochondria in the absence of precursor protein. Thus reticulocyte lysate contains an NEM‐sensitive, import stimulating factor which interacts both with the surface of mitochondria and whose activity appears to be dependent upon the structure of the mature portion of the precursor.

The Mitochondrial Protein Import Apparatus

“…70-kd stress proteins are generally ascribed a role in modulating the conformational and aggregational state of proteins in an ATP-dependent manner (58)(59)(60), and they might thus be good candidates for factors that confer transport competence to precursor proteins. A further proteinaceous cytosolic factor that is Annual Reviews www.annualreviews.org/aronline inactivated by treatment with N-ethylmaleimide seems to participate in protein transport into mitochondria (57,61). On the other hand, purified precursor proteins could be imported in the absence of cytosolic cofactors (47,62), suggesting that the requirement for cofactors is not a general one.…”

Mitochondrial import: properties of precursor proteins

“…The above findings do not define to what extent tified in recitulocyte lysate (Randall and Shore 1989). Furmitochondrial precursors must unfold in the process of being ther, depletion of a subset of 70-kDa stress proteins in yeasr Biochem.…”

Cytosolic and mitochondrial surface factor-independent import of a synthetic peptide into mitochondria.

“…Stimulation of the import of precursor proteins into isolated mitochondria by soluble factors in reticulocyte lysate or yeast cytosol was reported from several laboratories using bacterially expressed (Murakami and Mori, 1990;Sheffield et al, 1990) or in vitro synthesized (Murakami et al, 1988;Ono and Tuboi, 1990a;Randall and Shore, 1989) precursor proteins. Ono and Tuboi (1988) reported that the import of the OAT peptide was also stimulated by cytosolic factors in reticulocyte lysate.…”

“…P and M in (C) denote the precursor and the mature form of adrenodoxin, respectively. Effects of cytosolic factors on the import of 1 1,3-45 and adrenodoxin precursor It has recently been reported from several laboratories that cytosolic factors stimulate the import of precursor proteins (Murakami et al, 1988;Randall and Shore, 1989;Murakami and Mori, 1990) and a synthetic extension peptide (Ono and Tuboi, 1988) into isolated mitochondria. We examined the effect of cytosolic factors in reticulocyte lysate on the import of 111.45.…”

“…Thus the import of 111-45 was not dependent on cytosolic factors possibly because it was already in an import competent state. According to Randall and Shore (1989), the truncation of the carboxy-terminal 73 amino acid residues of ornithine transcarbamylase (OTC) precursor resulted in the loss of the dependence of its import on cytosolic factors, suggesting that recognition by cytosolic factors is not restricted to the extension peptide portion of the precursor. On the contrary, Murakami and Mori (1990) showed quite recently that presequence binding factor (PBF) which formed a soluble complex with OTC precursor was purified from reticulocyte lysate, and the formation of PBF -OTC precursor complex was inhibited by synthetic presequences (extension peptides) of OTC and other precursors.…”