Eukaryotic Mitochondria Research Articles

Targeting, Import, and Dimerization of a Mammalian Mitochondrial ATP Binding Cassette (ABC) Transporter, ABCB10 (ABC-me)

ATP binding cassette (ABC) transporters are a diverse superfamily of energy-dependent membrane translocases. Although responsible for the majority of transmembrane transport in bacteria, they are relatively uncommon in eukaryotic mitochondria. Organellar trafficking and import, in addition to quaternary structure assembly, of mitochondrial ABC transporters is poorly understood and may offer explanations for the paucity of their diversity. Here we examine these processes in ABCB10 (ABC-me), a mitochondrial inner membrane erythroid transporter involved in heme biosynthesis. We report that ABCB10 possesses an unusually long 105-amino acid mitochondrial targeting presequence (mTP). The central subdomain of the mTP (amino acids (aa) 36-70) is sufficient for mitochondrial import of enhanced green fluorescent protein. The N-terminal subdomain (aa 1-35) of the mTP, although not necessary for the trafficking of ABCB10 to mitochondria, participates in the proper import of the molecule into the inner membrane. We performed a series of amino acid mutations aimed at changing specific properties of the mTP. The mTP requires neither arginine residues nor predictable alpha-helices for efficient mitochondrial targeting. Disruption of its hydrophobic character by the mutation L46Q/I47Q, however, greatly diminishes its efficacy. This mutation can be rescued by cryptic downstream (aa 106-715) mitochondrial targeting signals, highlighting the redundancy of this protein's targeting qualities. Mass spectrometry analysis of chemically cross-linked, immunoprecipitated ABCB10 indicates that ABCB10 embedded in the mitochondrial inner membrane homodimerizes and homo-oligomerizes. A deletion mutant of ABCB10 that lacks its mTP efficiently targets to the endoplasmic reticulum. Quaternary structure assembly of ABCB10 in the ER appears to be similar to that in the mitochondria.

CpnDB: a chaperonin sequence database.

Type I chaperonins are molecular chaperones present in virtually all bacteria, some archaea and the plastids and mitochondria of eukaryotes. Sequences of cpn60 genes, encoding 60-kDa chaperonin protein subunits (CPN60, also known as GroEL or HSP60), are useful for phylogenetic studies and as targets for detection and identification of organisms. Conveniently, a 549-567-bp segment of the cpn60 coding region can be amplified with universal PCR primers. Here, we introduce cpnDB, a curated collection of cpn60 sequence data collected from public databases or generated by a network of collaborators exploiting the cpn60 target in clinical, phylogenetic, and microbial ecology studies. The growing database currently contains approximately 2000 records covering over 240 genera of bacteria, eukaryotes, and archaea. The database also contains over 60 sequences for the archaeal Type II chaperonin (thermosome, a homolog of eukaryotic cytoplasmic chaperonin) from 19 archaeal genera. As the largest curated collection of sequences available for a protein-encoding gene, cpnDB provides a resource for researchers interested in exploiting the power of cpn60 as a diagnostic or as a target for phylogenetic or microbial ecology studies, as well as those interested in broader subjects such as lateral gene transfer and codon usage. We built cpnDB from open source tools and it is available at http://cpndb.cbr.nrc.ca.

Bacterial proteins predisposed for targeting to mitochondria.

Mitochondria evolved from an endosymbiotic proteobacterium in a process that required the transfer of genes from the bacterium to the host cell nucleus, and the translocation of proteins thereby made in the host cell cytosol into the internal compartments of the organelle. According to current models for this evolution, two highly improbable events are required to occur simultaneously: creation of a protein translocation machinery to import proteins back into the endosymbiont and creation of targeting sequences on the protein substrates themselves. Using a combination of two independent prediction methods, validated through tests on simulated genomes, we show that at least 5% of proteins encoded by an extant proteobacterium are predisposed for targeting to mitochondria, and propose we that mitochondrial targeting information was preexisting for many proteins of the endosymbiont. We analyzed a family of proteins whose members exist both in bacteria and in mitochondria of eukaryotes and show that the amino-terminal extensions occasionally found in bacterial family members can function as a crude import sequence when the protein is presented to isolated mitochondria. This activity leaves the development of a primitive translocation channel in the outer membrane of the endosymbiont as a single hurdle to initiating the evolution of mitochondria.

Benzodiazepine binding to mitochondrial membranes of the amoeba Acanthamoeba castellanii and the yeast Saccharomyces cerevisiae.

Benzodiazepine binding sites were studied in mitochondria of unicellular eukaryotes, the amoeba Acathamoeba castellanii and the yeast Saccharomyces cerevisiae, and also in rat liver mitochondria as a control. For that purpose we applied Ro5-4864, a well-known ligand of the mitochondrial benzodiazepine receptor (MBR) present in mammalian mitochondria. The levels of specific [(3)H]Ro5-4864 binding, the dissociation constant (K(D)) and the number of [(3)H]Ro5-4864 binding sites (B(max)) determined for fractions of the studied mitochondria indicate the presence of specific [(3)H]Ro5-4864 binding sites in the outer membrane of yeast and amoeba mitochondria as well as in yeast mitoplasts. Thus, A. castellanii and S. cerevisiae mitochondria, like rat liver mitochondria, contain proteins able to bind specifically [(3)H]Ro5-4864. Labeling of amoeba, yeast and rat liver mitochondria with [(3)H]Ro5-4864 revealed proteins identified as the voltage dependent anion selective channel (VDAC) in the outer membrane and adenine nucleotide translocase (ANT) in the inner membrane. Therefore, the specific MBR ligand binding is not confined only to mammalian mitochondria and is more widespread within the eukaryotic world. However, it can not be excluded that MBR ligand binding sites are exploited efficiently only by higher multicellular eukaryotes. Nevertheless, the MBR ligand binding sites in mitochondria of lower eukaryotes can be applied as useful models in studies on mammalian MBR.

Control of redox transitions and oxygen species binding in Mn centers by biologically significant ligands; model studies with [Mn]-bacteriochlorophyll a.

Mn-superoxide dismutase (Mn-SOD), which protects the cell from the toxic potential of superoxide radicals (O(2)(-*)), is the only type of SOD which resides in eukaryotic mitochondria. Up-to-date, the exact catalytic mechanism of the enzyme and the relationship between substrate moieties and the ligands within the active site microenvironment are still not resolved. Here, we set out to explore the possible involvement of hydroperoxyl radicals ((*)OOH) in the catalytic dismutaion by following the interplay of Mn(III)/Mn(II) redox transitions, ligands binding, and evolution or consumption of superoxide radical, using a new model system. The model system encompassed an Mn atom chelated by a bacteriochlorophyll allomer macrocycle (BChl) in aerated aprotic media that contain residual water. The redox states of the Mn ion were monitored by the Q(y) electronic transitions at 774 and 825 nm for [Mn(II)]- and [Mn(III)]-BChl, respectively (Geskes, C.; Hartwich, G.; Scheer, H.; Mantele, W.; Heinze, J. J. Am. Chem. Soc. 1995, 117, 7776) and confirmed by electron spin resonance spectroscopy. Evolution of (*)OOH radicals was monitored by the ESR spin-trap technique using 5,5-dimethyl-1-pyrroline-N-oxide (DMPO). The experimental data suggest that the [Mn]-BChl forms a (HO(-))[Mn(III)]-BChl(OOH) complex upon solvation. Spectrophotometeric titrations with tetrabutylamonnium acetate (TBAA) and 1-methylimidazole (1-MeIm) together with ESI-MS measurements indicated the formation of a 1:1 complex with [Mn]-BChl for both ligands. The coordination of ligands at low concentrations to [Mn(III)]-BChl induced a release of a (*)OOH radical and a [Mn(III)]-BChl --> [Mn(II)]-BChl transition at higher concentrations. The estimated equilibrium constants for the total redox reaction ( )()are 1.9 x 10(4) +/- 1 x 10(3) M(-)(1) and 12.3 +/- 0.6 M(-)(1) for TBAA and 1-MeIm, respectively. The profound difference between the equilibrium constants agrees with the suggested key role of the ligand's basicity in the process. A direct interaction of superoxide radicals with [Mn(III)]-BChl in a KO(2) acetonitrile (AN) solution also resulted in [Mn(III)]-BChl --> [Mn(II)]-BChl transition. Cumulatively, our data show that the Mn(III) center encourages the protonation of the O(2)(-)(*) radical in an aprotic environment containing residual water molecules, while promoting its oxidation in the presence of basic ligands. Similar coordination and stabilization of the (*)OOH radical by the Mn center may be key steps in the enzymatic dismutation of superoxide radicals by Mn-SOD.

Biogenesis of iron–sulfur proteins in eukaryotes: components, mechanism and pathology

Iron–sulfur (Fe–S) clusters are ubiquitous co-factors of proteins that play an important role in metabolism, electron-transfer and regulation of gene expression. In eukaryotes mitochondria are the primary site of Fe–S cluster biogenesis. The organelles contain some ten proteins of the so-called iron–sulfur cluster (ISC) assembly machinery that is well-conserved in bacteria and eukaryotes. The ISC assembly machinery is responsible for biogenesis of Fe–S proteins within mitochondria. In addition, this machinery is involved in the maturation of extra-mitochondrial Fe–S proteins by cooperating with mitochondrial proteins with an exclusive function in this process. This review summarizes recent developments in our understanding of the biogenesis of cellular Fe–S proteins in eukaryotes. Particular emphasis is given to disorders in Fe–S protein biogenesis causing human disease.

Mitochondrial NAD-dependent methylenetetrahydrofolate dehydrogenase-methenyltetrahydrofolate cyclohydrolase is essential for embryonic development.

Folate-dependent enzymes are compartmentalized between the cytoplasm and mitochondria of eukaryotes. The role of mitochondrial folate-dependent metabolism and the extent of its contribution to cytoplasmic processes are areas of active investigation. NAD-dependent methylenetetrahydrofolate dehydrogenase-methenyltetrahydrofolate cyclohydrolase (NMDMC) catalyzes the interconversion of 5,10-methylenetetrahydrofolate and 10-formyltetrahydrofolate in mitochondria of mammalian cells, but its metabolic role is not yet clear. Its expression in embryonic tissues but not in most adult tissues as well as its stringent transcriptional regulation led us to postulate that it may play a role in embryonic development. To investigate the metabolic role of NMDMC, we used a knockout approach to delete the nmdmc gene in mice. Heterozygous mice appear healthy, but homozygous NMDMC knockout mice die in utero. At embryonic day 12.5 (E12.5), homozygous null embryos exhibit no obvious developmental defects but are smaller and pale and die soon thereafter. Mutant fetal livers contain fewer nucleated cells and lack the characteristic redness of wild-type or heterozygous livers. The frequencies of CFU-erythroid (CFU-E) and burst-forming unit-erythroid (BFU-E) from fetal livers of E12.5 null mutants were not reduced compared with those of wild-type or heterozygous embryos. It has been assumed that initiation of protein synthesis in mitochondria requires a formylated methionyl-tRNA(fmet). One role postulated for NMDMC is to provide 10-formyltetrahydrofolate as a formyl group donor for the synthesis of this formylmethionyl-tRNA(fmet). To determine if the loss of NMDMC impairs protein synthesis and thus could be a cause of embryonic lethality, mitochondrial translation products were examined in cells in culture. Mitochondrial protein synthesis was unaffected in NMDMC-null mutant cell lines compared with the wild type. These results show that NMDMC is not required to support initiation of protein synthesis in mitochondria in isolated cells but instead demonstrate an essential role for mitochondrial folate metabolism during embryonic development.

Recurrent intragenomic recombination leading to sequence homogenization during the evolution of the lipoyl-binding domain

The lipoyl-binding domain is often present, in one or several copies, in the E2 subunit and, less often, in the E1 and E3 subunits of 2-oxo acid dehydrogenase complexes. Phylogenetic analysis shows evidence of multiple, independent intragenomic recombination events between different versions of the lipoyl-binding domain in various bacteria and eukaryotic mitochondria, leading to homogenization of the sequences of the lipoyl-binding domain within the same enzymatic complex in several bacterial lineages. This appears to be the first case of sequence homogenization at the level of an individual domain in prokaryotes.

Characterization of isocitrate dehydrogenase from the green sulfur bacterium Chlorobium limicola. A carbon dioxide-fixing enzyme in the reductive tricarboxylic acid cycle

Isocitrate dehydrogenase (IDH) catalyzes the reversible conversion between isocitrate and 2-oxoglutarate accompanied by decarboxylation/carboxylation and oxidoreduction of NAD(P)+ cofactor. While this enzyme has been well studied as a catabolic enzyme in the tricarboxylic acid (TCA) cycle, here we have characterized NADP-dependent IDH from Chlorobium limicola, a green sulfur bacterium that fixes CO2 through the reductive tricarboxylic acid (RTCA) cycle, focusing on the CO2-fixation ability of the enzyme. The gene encoding Cl-IDH consisted of 2226 bp, corresponding to a polypeptide of 742 amino acid residues. The primary structure and the size of the recombinant protein indicated that Cl-IDH was a monomeric enzyme of 80 kDa distinct from the dimeric NADP-dependent IDHs predominantly found in bacteria or eukaryotic mitochondria. Apparent Michaelis constants for isocitrate (45 ± 13 µm) and NADP+ (27 ± 10 µm) were much smaller than those for 2-oxoglutarate (1.1 ± 0.5 mm) and CO2 (1.3 ± 0.3 mm). No significant differences in kinetic properties were observed between Cl-IDH and the dimeric, NADP-dependent IDH from Saccharomyces cerevisiae (Sc-IDH) at the optimum pH of each enzyme. However, in contrast to the 20% activity of Sc-IDH toward carboxylation as compared with that toward decarboxylation at pH 7.0, the activities of Cl-IDH for both directions were almost equivalent at this pH, suggesting a more favorable property of Cl-IDH than Sc-IDH as a CO2-fixation enzyme under physiological pH. Furthermore, we found that among various intermediates, oxaloacetate was a competitive inhibitor (Ki = 0.35 ± 0.04 mm) for 2-oxoglutarate in the carboxylation reaction by Cl-IDH, a feature not found in Sc-IDH.

Recurrent intragenomic recombination leading to sequence homogenization during the evolution of the lipoyl-binding domain

The lipoyl-binding domain is often present, in one or several copies, in the E2 subunit and, less often, in the E1 and E3 subunits of 2-oxo acid dehydrogenase complexes. Phylogenetic analysis shows evidence of multiple, independent intragenomic recombination events between different versions of the lipoyl-binding domain in various bacteria and eukaryotic mitochondria, leading to homogenization of the sequences of the lipoyl-binding domain within the same enzymatic complex in several bacterial lineages. This appears to be the first case of sequence homogenization at the level of an individual domain in prokaryotes.

On bacterial origin of mitochondria in eukaryotes in the light of current ideas of evolution of the organic world

The hypothesis of bacterial origin of mitochondria, which existed until the end of the 20th century, has been confirmed on the basis of the current concepts of organic world evolution in the open sea hydrosphere and original data on the entry of bacteria (prokaryotes0 in the cells of eukaryotes and their transformation into the mitochondrial mechanism of aerobic energy metabolism. This hypothesis can now be considered as a factually substantiated theory. The process of endocytosis of bacteria in the tissues of eukaryotes, which began at the onset of transition of the anaerobic state of open sea hydrosphere and land atmosphere (Early Proterozoic), is considered as the beginning of symbiotic mode of life of organisms of the Proterozoic and Postproterozoic organic world.

Mitochondrial type iron-sulfur cluster assembly in the amitochondriate eukaryotes Trichomonas vaginalis and Giardia intestinalis, as indicated by the phylogeny of IscS.

Pyridoxal-5'-phosphate-dependent cysteine desulfurase (IscS) is an essential enzyme in the assembly of FeS clusters in bacteria as well as in the mitochondria of eukaryotes. Although FeS proteins are particularly important for the energy metabolism of amitochondrial anaerobic eukaryotes, there is no information about FeS cluster formation in these organisms. We identified and sequenced two IscS homologs of Trichomonas vaginalis (TviscS-1 and TviscS-2) and one of Giardia intestinalis (GiiscS). TviscS-1, TviscS-2, and GiiscS possess the typical conserved regions implicated in cysteine desulfurase activity. N-termini of TviscS-1 and TviscS-2 possess eight amino acid extensions, which resemble the N-terminal presequences that target proteins to hydrogenosomes in trichomonads. No presequence was evident in GiiscS from Giardia, an organism that apparently lacks hydrogenosmes or mitochondria. Phylogenetic analysis showed a close relationship among all eukaryotic IscS genes including those of amitochondriates. IscS of proteobacteria formed a sister group to the eukaryotic clade, suggesting that isc-related genes were present in the proteobacterial endosymbiotic ancestor of mitochondria and hydrogenosomes. NifS genes of nitrogen-fixing bacteria, which are IscS homologs required for specific formation of FeS clusters in nitrogenase, formed a more distant group. The phylogeny indicates the presence of a common mechanism for FeS cluster formation in mitochondriates as well as in amitochondriate eukaryotes. Furthermore, the analyses support a common origin of Trichomonas hydrogenosomes and mitochondria, as well as secondary loss of mitochondrion/hydrogenosome-like organelles in Giardia.

Cloning and Characterization of Human and Mouse Mitochondrial Elongation Factor G, GFM and Gfm, and Mapping of GFM to Human Chromosome 3q25.1–q26.2

Similar to the translational system in the cell cytoplasm, the initiation, elongation, and termination of protein synthesis in the mitochondria of eukaryotes are catalyzed by several protein factors. These factors, from the viewpoint of evolution, are more closely related to the corresponding prokaryotic factors than to those in the eukaryotic cytoplasm. In this paper, we isolated two cDNAs coding for human and mouse mitochondrial elongation factor G (GFM and Gfm, respectively). The GFM cDNA, which is 3481 bp in length, predicts a protein of 751 amino acids sharing 84 and 42% identity and 88 and 62% similarity to rat EF-Gmt and Escherichia coli EF-G, respectively, and 24% identity and 39% similarity to human EF-2, the equivalent of EF-G in the cytoplasm. The mouse Gfm cDNA is 2564 bp and contains an intact open reading frame that encodes 751 amino acids showing 89% sequence identity and 94% similarity to human GFM. Northern blot analysis of human GFM revealed three transcripts of 3.8, 3.4, and 2.9 kb. The first two were expressed at high levels in heart, skeletal muscle, and testis, at moderate levels in liver and kidney, and at low levels in other tissues including brain, placenta, and lung, while the last transcript was expressed only in testis. The relative abundance of GFM was consistent with the observations for human EF-Tumt and EF-Tsmt, the other two mitochondrial elongation factors, indicating that the three factors were expressed at corresponding levels. The expression pattern of mouse Gfm was also determined, which showed that Gfm was expressed as a 3.0-kb transcript, abundantly in heart, skeletal muscle, kidney, and testis. In addition, GFM was assigned to human chromosome 3q25.1–q26.2 by the radiation hybrid mapping method. The genomic organization of GFM was also analyzed by comparing this cDNA with a genomic DNA sequence (Accession No. AC010936), which showed that GFM contained 18 exons and spanned at least 40 kb.

Molecular evolution of the AMP-forming Acetyl-CoA synthetase

Acetyl-CoA-Synthetase (ACS) is involved in the production of acetate, a major metabolite in numerous organisms. There are two forms of this enzyme: ADP-forming ACS and ATP-forming ACS. We focus mainly on the AMP-forming ACS gene, which is relatively well conserved in eubacteria, archeaebacteria, and eukaryotes. BLAST searches in databases showed 30 protein sequences significantly related to the ACS. Most of these sequences were identified as ACS but three of them, belonging to the mammalian species, were annotated as another gene named: the SA gene, which is involved in the essential hypertension. The ACS and SA genes probably derived from a duplication of an ancestral gene but have acquired different functions. Six conserved regions of the ACS protein were defined across the three domains of life. While the precise function of the conserved regions remains unknown, they are probably involved in the enzymatic activity. Among eukaryotes, we found a high variability with respect to the number and the position of introns. However, some positions are conserved between fungi and a nematode. A maximum likelihood tree based upon the conserved regions showed that all sequences except the one from B.subtilis, belong to two basic groups: one the SA-like group including sequences from Archaeoglobusfulgidus and Streptomycescoelicolor, and second, the ACS group. The later can be further divided in two parts: a prokaryotic one including eubacteria and an archaebacterium, and a eukaryotic group within which two proteobacterial sequences branch including ACS from the α-proteobacterium Rhodobactercapsulatus. Within the eukaryotic group, bootstrap support is very low, but overall the data are consistent with the view that eukaryotes acquired their ACS gene from the ancestors of mitochondria. The localization of this enzyme in eukaryotic mitochondria is the additional evidence in favor of this interpretation.

The Energy Significance of Symbiotic Bacteria in the Metabolism of Hydrothermal and Other Chemotrophic Biota of the Open Sea

The bacterial origin of eukaryotic mitochondria, specifically in Metazoa, as a mechanism of their basic (aerobic) respiration and the role of symbiotic bacteria during the supply of energy to the metazoan host is proved for the first time from the viewpoint of the monophyletic development of the organic world and the origin of eukaryotes as descendants of prokaryotes. Representatives of the hydrothermal bacteriochemosymbiotrophic bottom fauna of the open sea were used as examples.

Four genes are required for the system II cytochrome c biogenesis pathway in Bordetella pertussis, a unique bacterial model.

Unlike other cytochromes, c-type cytochromes have two covalent bonds formed between the two vinyl groups of haem and two cysteines of the protein. This haem ligation requires specific assembly proteins in prokaryotes or eukaryotic mitochondria and chloroplasts. Here, it is shown that Bordetella pertussis is an excellent bacterial model for the widespread system II cytochrome c synthesis pathway. Mutations in four different genes (ccsA, ccsB, ccsX and dipZ) result in B. pertussis strains unable to synthesize any of at least seven c-type cytochromes. Using a cytochrome c4:alkaline phosphatase fusion protein as a bifunctional reporter, it was demonstrated that the B. pertussis wild-type and mutant strains secrete an active alkaline phosphatase fusion protein. However, unlike the wild type, all four mutants are unable to attach haem covalently, resulting in a degraded N-terminal apocytochrome c4 component. Thus, apocytochrome c secretion is normal in each of the four mutants, but all are defective in a periplasmic assembly step (or export of haem). CcsX is related to thioredoxins, which possess a conserved CysXxxXxxCys motif. Using phoA gene fusions as reporters, CcsX was proven to be a periplasmic thioredoxin-like protein. Both the B. pertussis dipZ (i. e. dsbD) and ccsX mutants are corrected for their assembly defects by the thiol-reducing compounds, dithiothreitol and 2-mercaptoethanesulphonic acid. These results indicate that DipZ and CcsX are required for the periplasmic reduction of the cysteines of apocytochromes c before ligation. In contrast, the ccsA and ccsB mutants are not corrected by exogenous reducing agents, suggesting that CcsA and CcsB are required for the haem ligation step itself in the periplasm (or export of haem to the periplasm). Related to this suggestion, the topology of CcsB was determined experimentally, demonstrating that CcsB has four transmembrane domains and a large 435-amino-acid periplasmic region.

The respiratory complex I of bacteria, archaea and eukarya and its module common with membrane-bound multisubunit hydrogenases

The proton-pumping NADH:ubiquinone oxidoreductase, also called complex I, is the first of the respiratory complexes providing the proton motive force which is essential for energy consuming processes like the synthesis of ATP. Homologues of this complex exist in bacteria, archaea, in mitochondria of eukaryotes and in chloroplasts of plants. The bacterial and mitochondrial complexes function as NADH dehydrogenase, while the archaeal complex works as F 420H 2 dehydrogenase. The electron donor of the cyanobacterial and plastidal complex is not yet known. Despite the different electron input sites, 11 polypeptides constitute the structural framework for proton translocation and quinone binding in the complex of all three domains of life. Six of them are also present in a family of membrane-bound multisubunit [NiFe] hydrogenases. It is discussed that they build a module for electron transfer coupled to proton translocation.

Crystallization and preliminary X-ray diffraction analysis of the mitochondrial transcription factor sc-mtTFB from Saccharomyces cerevisiae.

Eukaryotic mitochondria contain a distinct mini-chromosome. In yeast, transcription of the mitochondrial genome is mediated by a nuclear-encoded RNA polymerase consisting of a single polypeptide core enzyme and a specificity factor termed sc-mtTFB which bears some similarity to bacterial sigma-factors. sc-mtTFB from Saccharomyces cerevisiae has been cloned, expressed, purified and crystallized. The crystals belong to the monoclinic space group C2, with unit-cell parameters a = 89.7, b = 44.6, c = 98.9 A, beta = 110 degrees. Based on one molecule per asymmetric unit, the solvent content is estimated to be 48%. Small crystals of dimensions 0.01 x 0.05 x 0.13 mm diffract to at least 2.7 A resolution on a rotating-anode X-ray source.

What is the role of epsilon in the Escherichia coli ATP synthase?

The ATP synthase from Escherichia coli is a prototype of the ATP synthases that are found in many bacteria, in the mitochondria of eukaryotes, and in the chloroplasts of plants. It contains eight different types of subunits that have traditionally been divided into F(1), a water-soluble catalytic sector, and F(o), a membrane-bound ion transporting sector. In the current rotary model for ATP synthesis, the subunits can be divided into rotor and stator subunits. Several lines of evidence indicate that epsilon is one of the three rotor subunits, which rotate through 360 degrees. The three-dimensional structure of epsilon is known and its interactions with other subunits have been explored by several approaches. In light of recent work by our group and that of others, the role of epsilon in the ATP synthase from E. coli is discussed.

The Heme Biosynthesis Pathway and Clinical Manifestations of Abnormal Function

Biosynthesis of heme is important for both prokaryotes and eukaryotes. The enzymes for this multistep process are distributed between the cytosol and mitochondria in eukaryotes. In humans there are inherited and acquired disorders characterized by over synthesis of one or more enzymes or absence of an enzyme. This overview discusses each enzyme in the heme biosynthesis pathway.