The mitochondrial targeting function of randomly generated peptide sequences correlates with predicted helical amphiphilicity.

A pool of oligonucleotides encoding a start methionine and nine random amino acids was inserted at the 5'-end of the gene for the yeast cytochrome oxidase subunit IV lacking its own mitochondrial targeting sequence. Approximately one-quarter of the randomly generated sequences targeted subunit IV to its correct intramitochondrial location in vivo. Sequence analysis of 89 randomly generated sequences showed that their efficiencies as mitochondrial targeting signals correlated with the potential to fold into an amphiphilic alpha-helix. Functional targeting sequences were enriched in arginine and isoleucine residues but contained few aspartate, glutamate, and proline residues. Nonfunctional sequences predicted to have significant helical amphiphilicity often had at least one acidic or multiple helix-breaking residues that would be expected to interfere with targeting functioning. These results support the hypothesis that the signal for targeting a protein into the mitochondrial matrix is usually a positively charged amphiphilic helix.

Postglacial recolonization at a snail's pace (Trochulus villosus): confronting competing refugia hypotheses using model selection.

The localization of Last Glacial Maximum (LGM) refugia is crucial information to understand a species' history and predict its reaction to future climate changes. However, many phylogeographical studies often lack sampling designs intensive enough to precisely localize these refugia. The hairy land snail Trochulus villosus has a small range centred on Switzerland, which could be intensively covered by sampling 455 individuals from 52 populations. Based on mitochondrial DNA sequences (COI and 16S), we identified two divergent lineages with distinct geographical distributions. Bayesian skyline plots suggested that both lineages expanded at the end of the LGM. To find where the origin populations were located, we applied the principles of ancestral character reconstruction and identified a candidate refugium for each mtDNA lineage: the French Jura and Central Switzerland, both ice-free during the LGM. Additional refugia, however, could not be excluded, as suggested by the microsatellite analysis of a population subset. Modelling the LGM niche of T. villosus, we showed that suitable climatic conditions were expected in the inferred refugia, but potentially also in the nunataks of the alpine ice shield. In a model selection approach, we compared several alternative recolonization scenarios by estimating the Akaike information criterion for their respective maximum-likelihood migration rates. The 'two refugia' scenario received by far the best support given the distribution of genetic diversity in T. villosus populations. Provided that fine-scale sampling designs and various analytical approaches are combined, it is possible to refine our necessary understanding of species responses to environmental changes.

COX24 codes for a mitochondrial protein required for processing of the COX1 transcript

In most strains of Saccharomyces cerevisiae the mitochondrial gene COX1, for subunit 1 of cytochrome oxidase, contains multiple exons and introns. Processing of COX1 primary transcript requires accessory proteins factors, some of which are encoded by nuclear genes and others by reading frames residing in some of the introns of the COX1 and COB genes. Here we show that the low molecular weight protein product of open reading frame YLR204W, for which we propose the name COX24, is also involved in processing of COX1 RNA intermediates. The growth defect of cox24 mutants is partially rescued in strains harboring mitochondrial DNA lacking introns. Northern blot analyses of mitochondrial transcripts indicate cox24 null mutants to be blocked in processing of introns aI2 and aI3. The dependence of intron aI3 excision on Cox24p is also supported by the growth properties of the cox24 mutant harboring mitochondrial DNA with different intron compositions. The intermediate phenotype of the cox24 mutant in the background of intronless mitochondrial DNA, however, suggests that in addition to its role in splicing of the COX1 pre-mRNA, Cox24p still has another function. Based on the analysis of a cox14-cox24 double mutant, we propose that the other function of Cox24p is related to translation of the COX1 mRNA. © 2006 by The American Society for Biochemistry and Molecular Biology, Inc.

Nuclear mitochondrial-like sequences in ants: Evidence from Atta cephalotes (Formicidae: Attini)

Nuclear mitochondrial-like sequences (numts) are copies of mitochondrial DNA that have migrated to the genomic DNA. We present the first characterization of numts in ants, these numts being homologues to a mitochondrial DNA fragment containing loci the 3′ portion of the cytochrome oxidase I gene, an intergenic spacer, the tRNA leucine gene and the 5′ portion of the cytochrome oxidase II gene. All 67 specimens of Atta cephalotes (Hymenoptera: Formicidae: Attini) investigated had these homologues, which are within two monophyletic groups that we called numt1 and numt2. Numt1 and numt2 sequences are less variable than mitochondrial sequences and released from the severe purifying selection constraining the evolution of mitochondrial genes. Their formation probably involved bottlenecks related to two distinct transfer events of ancient and fast evolving mitochondrial DNA fragments to comparative slowly evolving nuclear DNA regions. © 2007 The Authors.

Translational regulation of nuclear gene COX4 expression by mitochondrial content of phosphatidylglycerol and cardiolipin in Saccharomyces cerevisiae.

Previous results indicated that translation of four mitochondrion-encoded genes and one nucleus-encoded gene (COX4) is repressed in mutants (pgs1Delta) of Saccharomyces cerevisiae lacking phosphatidylglycerol and cardiolipin. COX4 translation was studied here using a mitochondrially targeted green fluorescence protein (mtGFP) fused to the COX4 promoter and its 5' and 3' untranslated regions (UTRs). Lack of mtGFP expression independent of carbon source and strain background was established to be at the translational level. The translational defect was not due to deficiency of mitochondrial respiratory function but was rather caused directly by the lack of phosphatidylglycerol and cardiolipin in mitochondrial membranes. Reintroduction of a functional PGS1 gene under control of the ADH1 promoter restored phosphatidylglycerol synthesis and expression of mtGFP. Deletion analysis of the 5' UTR(COX4) revealed the presence of a 50-nucleotide fragment with two stem-loops as a cis-element inhibiting COX4 translation. Binding of a protein factor(s) specifically to this sequence was observed with cytoplasm from pgs1Delta but not PGS1 cells. Using HIS3 and lacZ as reporters, extragenic spontaneous recessive mutations that allowed expression of His3p and beta-galactosidase were isolated, which appeared to be loss-of-function mutations, suggesting that the genes mutated may encode the trans factors that bind to the cis element in pgs1Delta cells.

Comparison of two measures of PSI electron transport in winter rye

The number of P700 (the reaction centre of Photosystem I) converted to P700+, in winter rye, was determined by measuring the absorbance change at 820nm . It was found, with a single turnover flash, that thylakoids isolated from cold grown plants have a 50% greater number of P700 oxidized than thylakoids isolated from warm grown plants. Incubation of thylakoids in the dark at 35 C did not change the number of P700 oxidized. The conversion of P700 to P700+ with a single flash can be compared to a steady state rate of electron transport using a Clark electrode. The results for P700 oxidation using the absorbance change at 820 nm measure effects within the PSI complex whereas the results obtained from a Clark electrode measures steady state electron transport between the cytochrome blf complex and the PSI complex. In contrast to the results for P700 oxidation it was shown, using a Clark electrode, that both thylakoids from cold grown plants and thylakoids incubated at in the dark 35 C exhibited 50% higher rates of electron transport than thylakoids from warm grown plants. The correlation between the higher rate of steady state PSI electron transport observed in thylakoids isolated from cold grown winter rye and number of active PSI reaction centres localizes the site of the increase to the PSI reaction centre. In contrast the lack of correlation after incubation at 35 C indicates the increase in the rate of light saturated electron transport in thylakoids isolated from cold grown plants and thylakoids incubated in the dark at 35 C occur by different mechanisms.

Palmitate increases superoxide production through mitochondrial electron transport chain and NADPH oxidase activity in skeletal muscle cells

The effect of unbound palmitic acid (PA) at plasma physiological concentration range on reactive oxygen species (ROS) production by cultured rat skeletal muscle cells was investigated. The participation of the main sites of ROS production was also examined. Production of ROS was evaluated by cytochrome c reduction and dihydroethidium oxidation assays. PA increased ROS production after 1 h incubation. A xanthine oxidase inhibitor did not change PA-induced ROS production. However, the treatment with a mitochondrial uncoupler and mitochondrial complex III inhibitor decreased superoxide production induced by PA. The importance of mitochondria was also evaluated in 1 h incubated rat soleus and extensor digitorum longus (EDL) muscles. Soleus muscle, which has a greater number of mitochondria than EDL, showed a higher superoxide production induced by PA. These results indicate that mitochondrial electron transport chain is an important contributor for superoxide formation induced by PA in skeletal muscle. Results obtained with etomoxir and bromopalmitate treatment indicate that PA has to be oxidized to raise ROS production. A partial inhibition of superoxide formation induced by PA was observed by treatment with diphenylene iodonium, an inhibitor of NADPH oxidase. The participation of this enzyme complex was confirmed through an increase of p47(phox) phosphorylation after treatment with PA.

A dedicated setup for the measurement of the electron transport parameters in gases at large electric fields

Electron transport parameters are important in several areas ranging from particle detectors to plasma-assisted processing reactors. Nevertheless, especially at high fields strengths and for complex gases, relatively few data are published. A dedicated setup has been developed to measure the electron drift velocity and the first Townsend coefficient in parallel plate geometry. An RPC-like cell has been adopted to reach high field strengths without the risk of destructive sparks. The validation data obtained with pure Nitrogen will be presented and compared to a selection of the available literature and to calculations performed with Magboltz 2 version 8.6. The new data collected in pure Isobutane will then be discussed. This is the first time the electron drift velocity in pure Isobutane is measured well into the saturation region. Good agreement is found with expectations from Magboltz. (C) 2009 Elsevier B.V. All rights reserved.

Development of a fluorescent method for the detection in marine organisms of the respiratory electron transport system

Trabajo realizado por: Maldonado, F.; Packard, T.; Gómez, M.; Santana Rodríguez, J. J

Association of glycolate oxidation with photosynthetic electron transport in plant and algal chloroplasts.

Photosynthetic carbon metabolism is initiated by ribulose-bisphosphate carboxylase/oxygenase (Rubisco), which uses both CO2 and O2 as substrates. One 2-phosphoglycolate (P-glycolate) molecule is produced for each O2 molecule fixed. P-glycolate has been considered to be metabolized exclusively via the oxidative photosynthetic carbon cycle. This paper reports an additional pathway for P-glycolate and glycolate metabolism in the chloroplasts. Light-dependent glycolate or P-glycolate oxidation by osmotically shocked chloroplasts from the algae Dunaliella or spinach leaves was measured by three electron acceptors, methyl viologen (MV), potassium ferricyanide, or dichloroindophenol. Glycolate oxidation was assayed with 3-(3,4)-dichlorophenyl)-1,1-dimethylurea (DCMU) as oxygen uptake in the presence of MV at a rate of 9 mol per mg of chlorophyll per h. Washed thylakoids from spinach leaves oxidized glycolate at a rate of 22 mol per mg of chlorophyll per h. This light-dependent oxidation was inhibited completely by SHAM, an inhibitor of quinone oxidoreductase, and 75% by 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB), which inhibits electron transfer from plastoquinone to the cytochrome b6f complex. SHAM stimulated severalfold glycolate excretion by algal cells, Dunaliella or Chlamydomonas, and by isolated Dunaliella chloroplasts. Glycolate and P-glycolate were oxidized about equally well to glyoxylate and phosphate. On the basis of results of inhibitor action, the possible site which accepts electrons from glycolate or P-glycolate is a quinone after the DCMU site but before the DBMIB site. This glycolate oxidation is a light-dependent, SHAM-sensitive, glycolate-quinone oxidoreductase system that is associated with photosynthetic electron transport in the chloroplasts.

Improvement Of The Physical Performance Is Associated With Activation Of No/pgc-1α/mttfa Signaling Pathway And Increased Protein Expressions Of Electron Transport Chain In Gastrocnemius Muscle From Rats Supplemented With L-arginine.

To examine the influence of l-arginine supplementation in combination with physical training on mitochondrial biomarkers from gastrocnemius muscle and its relationship with physical performance. Male Wistar rats were divided into four groups: control sedentary (SD), sedentary supplemented with l-arginine (SDLA), trained (TR) and trained supplemented with l-arginine (TRLA). Supplementation of l-arginine was administered by gavage (62.5mg/ml/day/rat). Physical training consisted of 60min/day, 5days/week, 0% grade, speed of 1.2km/h. The study lasted 8weeks. Skeletal muscle mitochondrial enriched fraction as well as cytoplasmic fractions were obtained for Western blotting and biochemical analyses. Protein expressions of transcriptor coactivator (PGC-1α), transcriptor factors (mtTFA), ATP synthase subunit c, cytochrome oxidase (COXIV), constitutive nitric oxide synthases (eNOS and nNOS), Cu/Zn-superoxide dismutase (SOD) and manganese-SOD (Mn-SOD) were evaluated. We also assessed in plasma: lipid profile, glycemia and malondialdehyde (MDA) levels. The nitrite/nitrate (NOx(-)) levels were measured in both plasma and cytosol fraction of the gastrocnemius muscle. 8-week l-arginine supplementation associated with physical training was effective in promoting greater tolerance to exercise that was accompanied by up-regulation of the protein expressions of mtTFA, PGC-1α, ATP synthase subunit c, COXIV, Cu/Zn-SOD and Mn-SOD. The upstream pathway was associated with improvement of NO bioavailability, but not in NO production since no changes in nNOS or eNOS protein expressions were observed. This combination would be an alternative approach for preventing cardiometabolic diseases given that in overt diseases a profound impairment in the physical performance of the patients is observed.

An in situ study of photosynthetic oxygen exchange and electron transport rate in the marine macroalga Ulva lactuca (Chlorophyta)

Direct comparisons between photosynthetic O-2 evolution rate and electron transport rate (ETR) were made in situ over 24 h using the benthic macroalga Ulva lactuca (Chlorophyta), growing and measured at a depth of 1.8 m, where the midday irradiance rose to 400-600 mumol photons m(-2) s(-1). O-2 exchange was measured with a 5-chamber data-logging apparatus and ETR with a submersible pulse amplitude modulated (PAM) fluorometer (Diving-PAM). Steady-state quantum yield ((Fm'-Ft)/Fm') decreased from 0.7 during the morning to 0.45 at midday, followed by some recovery in the late afternoon. At low to medium irradiances (0-300 mumol photons m(-2) s(-1)), there was a significant correlation between O-2 evolution and ETR, but at higher irradiances, ETR continued to increase steadily, while O-2 evolution tended towards an asymptote. However at high irradiance levels (600-1200 mumol photons m-(2) s(-1)) ETR was significantly lowered. Two methods of measuring ETR, based on either diel ambient light levels and fluorescence yields or rapid light curves, gave similar results at low to moderate irradiance levels. Nutrient enrichment (increases in [NO3-], [NH4+] and [HPO42-] of 5- to 15-fold over ambient concentrations) resulted in an increase, within hours, in photosynthetic rates measured by both ETR and O-2 evolution techniques. At low irradiances, approximately 6.5 to 8.2 electrons passed through PS II during the evolution of one molecule of O-2, i.e., up to twice the theoretical minimum number of four. However, in nutrient-enriched treatments this ratio dropped to 5.1. The results indicate that PAM fluorescence can be used as a good indication of the photosynthetic rate only at low to medium irradiances.

The electron transfer complex between nitrous oxide reductase and its electron donors

J Biol Inorg Chem (2011) 16:1241–1254 DOI 10.1007/s00775-011-0812-9

Electron transfer complex between nitrous oxide reductase and cytochrome c552 from Pseudomonas nautica: kinetic, nuclear magnetic resonance, and docking studies

Biochemistry. 2008 Oct 14;47(41):10852-62. doi: 10.1021/bi801375q

Monte Carlo simulation of kilovolt electron transport in solids

A Monte Carlo procedure to simulate the penetration and energy loss of low¿energy electron beams through solids is presented. Elastic collisions are described by using the method of partial waves for the screened Coulomb field of the nucleus. The atomic charge density is approximated by an analytical expression with parameters determined from the Dirac¿Hartree¿Fock¿Slater self¿consistent density obtained under Wigner¿Seitz boundary conditions in order to account for solid¿state effects; exchange effects are also accounted for by an energy¿dependent local correction. Elastic differential cross sections are then easily computed by combining the WKB and Born approximations to evaluate the phase shifts. Inelastic collisions are treated on the basis of a generalized oscillator strength model which gives inelastic mean free paths and stopping powers in good agreement with experimental data. This scattering model is accurate in the energy range from a few hundred eV up to about 50 keV. The reliability of the simulation method is analyzed by comparing simulation results and experimental data from backscattering and transmission measurements.