Showing papers on "Mitochondrial carrier published in 1984"

The amino-terminal region of an imported mitochondrial precursor polypeptide can direct cytoplasmic dihydrofolate reductase into the mitochondrial matrix

Abstract: Subunit IV of yeast cytochrome c oxidase is encoded by a nuclear gene, synthesized in the cytosol as a precursor with a transient amino-terminal extension of 25 amino acids, and imported into the mitochondria. By gene fusion, we have attached the amino-terminal 53 amino acids of the subunit IV precursor to the amino terminus of the mouse cytosolic enzyme dihydrofolate reductase. When the resulting fusion protein was synthesized in a transcription-translation system and then incubated with energized yeast mitochondria, it was imported into the mitochondrial matrix space and processed to a shorter form by the chelator-sensitive matrix protease. No evidence was obtained that the fusion protein became stuck across one of the two mitochondrial membranes. Thus, a non-mitochondrial protein can be transported into the mitochondrial matrix if it is fitted with a mitochondrial targeting sequence.

A 70-kd protein of the yeast mitochondrial outer membrane is targeted and anchored via its extreme amino terminus

Abstract: The major 70-kd protein of the yeast mitochondrial outer membrane is made on cytosolic ribosomes and imported into the outer membrane without proteolytic cleavage. We have attempted to identify the sequences which target the protein to the mitochondria and which permanently anchor it to the lipid bilayer of the outer membrane. By manipulating the cloned gene we have deleted 13 different regions throughout the polypeptide; in addition, we have fused amino-terminal regions of different length to beta-galactosidase. Each altered gene was introduced into yeast and the intracellular fate of the corresponding polypeptide product was determined by subcellular fractionation. All the information for targeting and anchoring the 70-kd protein (617 amino acids) was contained within the amino-terminal 41 amino acids. When this entire region was deleted, the protein was recovered with the cytosol fraction. However, several restricted deletions within this amino-terminal region appeared to affect targeting and anchoring differentially: most of the altered protein remained in the cytosol but a small fraction was misrouted into the mitochondrial matrix space. We suggest that targeting is mediated by a region which includes the 11 amino-terminal amino acids whereas the permanent membrane anchor is provided by a typical transmembrane sequence between residues 9 and 38.

Chapter 8 Metabolite transport in mammalian mitochondria

Abstract: Publisher Summary The chapter offers information on metabolite transport in mammalian mitochondria. A steady flow of metabolites, both in and out of the mitochondrial matrix space is necessary for mitochondria to perform functions that involve the participation of enzymes inside the membrane permeability barrier. In some mammalian cells, enzymes comprising partial spans of biosynthetic pathways are inside and some outside the mitochondrial matrix space. Therefore, in the liver—six mitochondrial membrane transport proteins are required for urea synthesis, three for gluconeogenesis, and three others participate in ammoniagenesis in the kidney. Most metabolites that are substrates of mitochondrial transporters are anions at physiological pH. Inhibitors specific to some of the transporters are identified and these could be used to establish the participation of the transporters in metabolic pathways. The chapter presents the carriers that catalyze net uptake of ions in symport (or antiport) with protons that neutralize the anion's charge. These carriers include the phosphate carrier, the monocarboxylate carrier that catalyzes the transport of pyruvate, acetoacetate and possibly branched chainketoacids, and the glutamate carrier. A second category of carriers catalyzes 1: 1 electroneutral exchanges of anions across the membrane. A third classification includes those transporters that catalyze transport of metabolites that are neutral at physiological pH. These include the transporters for glutamine, and for the other neutral amino acids. The last category includes the electrogenic or, the electrophoretic transporters.

Biogenesis of mitochondria: Defective yeast H+-ATPase assembled in the absence of mitochondrial protein synthesis is membrane associated

Abstract: We have investigated the extent to which the assembly of the cytoplasmically synthesized subunits of the H+-ATPase can proceed in a mtDNA-less (rho degree) strain of yeast, which is not capable of mitochondrial protein synthesis. Three of the membrane sector proteins of the yeast H+-ATPase are synthesized in the mitochondria, and it is important to determine whether the presence of these subunits is essential for the assembly of the imported subunits to the inner mitochondrial membrane. A monoclonal antibody against the cytoplasmically synthesized beta-subunit of the H+-ATPase was used to immunoprecipitate the assembled subunits of the enzyme complex. Our results indicate that the imported subunits of the H+-ATPase can be assembled in this mutant, into a defective complex which could be shown to be associated with the mitochondrial membrane by the analysis of the Arrhenius kinetics of the mutant mitochondrial ATPase activity.

Transport and regulation of polypeptide precursors of mature mitochondrial proteins.

Abstract: Publisher Summary Most mitochondrial proteins are coded by nuclear genes, synthesized on cytosolic ribosomes, and then imported into mitochondria. As is the case for some other proteins, most mitochondrial proteins are synthesized as larger precursors. This chapter discusses some of the characteristics and regulatory aspects of protein transport from cytosol to mitochondria. The large majority of mitochondrial proteins are coded by genes of the nucleus, synthesized in the cytoplasmic ribosomes, and then transported to the mitochondria. This seems to be the type of synthesis that takes place for the proteins of the mitochondrial matrix, those of the outer membrane, and most of the proteins of the inner membrane. The enzymes that participate in the replication, transcription, and translation of mitochondrial DNA are among the proteins synthesized in this way. The most widely accepted explanation of why most mitochondrial proteins are synthesized in the cytosol despite having their own genetic system is related to the possible evolutionary origin of this organelle. In this respect, it has been demonstrated recently that in Neurospora crassa, there is a gene in the mitochondrial DNA that codes for a protein homologous to one coded for by nuclear DNA. This suggests and is congruent with the endosymbiotic hypothesis for the origin of eukaryotic mitochondria-gene duplication followed by the transfer of one of these genes to the nucleus. This transfer of DNA between organelles has been extended to the passage of DNA from the chloroplast to the mitochondria.

Transport of the Precursor for Sulfite Oxidase into Intermembrane Space of Liver Mitochondria: Binding of the Precursor to Outer Mitochondrial Membrane

Abstract: Rat liver mitochondria were incubated with in vitro translation products programmed by liver RNA, then disrupted by sonication and subjected to sucrose density gradient centrifugation. Pre-sulfite oxidase bound preferentially to the outer mitochondrial membrane recovered with the inner membrane. This outer membrane could not be separated from the inner membrane by recentrifugation, suggesting the tight association of both membranes. The binding was not affected by pretreatment of mitochondria with proteolytic enzymes. The mature enzyme and its precursor synthesized in isolated hepatocytes have isoelectric points of 4.2 and 5.5, respectively. The molecular size of the precursor in cytosol was estimated to be about 100,000 daltons (dimer) by gel filtration.

The mitochondrial carrier of adenylates controls ATP production in the physiological range of respiration rates

Abstract: The problem is considered concerning the amount of control exerted by different mitochondrial enzymes on oxidative phosphorylation. Using the data of Groen et al. (1982) it has been found that when the respiration rates for isolated mitochondria ranged from 30 to 50 per cent of that in state 3 (which is in apparent physiological range) the contribution of the adenine nucleotide translocator to the control of ATP production was no less than 90 per cent taking for 100 per cent the total contribution of all mitochondrial enzymes.

Transport of mature proteins into isolated mitochondria: a model system to investigate mitochondrial biogenesis.

Abstract: movement of AAT still occurred. Furthermore, phenylsuccinate, a non-permeant, non-metabolizable analogue of succinate also triggered externalization of the enzyme. (b) Translocation does not require the modification of the primary structure of ‘mobile’ AAT. After externalization, AAT was purified, labelled with I z 5 I and internalized into mitoplasts, after which it was reexternalized by addition of succinate. In these three situations (out, in, and out of the mitoplast), the molecular weight of AAT was compared by gel electrophoresis. The results clearly show no difference of apparent molecular weight. A configurational modification induced by the environment might thus be responsible for the reversible movement of protein. (c) The driving force for this protein movement may possibly be the chemical potential of the interacting membrane components and translocating protein as modulated by the presence or absence of movement effectors (specific molecules, salts, pH, etc.). This is best illustrated by the experiments of Furuya et al. (1979) in which it was shown that mitochondrial AAT becomes latent and nonaccessible to proteases, and thus probably internalized, when incubated in the presence of negatively charged liposomes but not in the presence of positively charged ones. As for the mechanisms involved in reversible or vectorial internalization of protein, a general mechanism has recently been proposed. It suggests that the primary event in insertion of proteins into and translocation through membranes could be the spontaneous hydrophobically driven partitioning of a helical hairpin into the membrane bilayer (Engelman & Steitz, 1981).