Intramolecular G-quadruplexes from microsatellite d(GT)12 sequence in the presence of K+

Polymorphic d(GT)n microsatellite sequences are known to drastically affect genes expression. By use of CD spectroscopy, UV melting, fluorescence polarization of EtBr probe and FRET, we detected formation of a new fold with three G-quartets by d(GT)12 oligonucleotide in 0.01 M Na phosphate buffer, pH 8.0, in the presence 0.1 M KCl. Monomolecular type of the structure was verified with measurements of rotational relaxation time (p = 28 +/- 0.5 ns) of EtBr:d(GT)12 complex. CD spectra supported G-quartets formation. A distance between FITC, covalently attached to 5'-end of d(GT)12, and intercalated EtBr molecule was estimated using FRET (R < or =17 A). These data are in agreement with the proposed self folding of d(GT)12.     

Influence of Zn<Superscript>2+</Superscript>, Co<Superscript>2+</Superscript> and dimethylbenzimidazole on vitamin B<Subscript>12</Subscript> biosynthesis by <Emphasis Type="Italic">Pseudomonas denitrificans</Emphasis>

Previous research has confirmed that cobalt ion and dimethylbenzimidazole (DMBI) are the precursors of vitamin B₁₂ biosynthesis, and porphobilinogen synthase (PBG synthase) is a zinc-requiring enzyme. In this paper, the effects of Zn²⁺, Co²⁺ and DMBI on vitamin B₁₂ production by Pseudomonas denitrificans in shake flasks were studied. Present experimental results demonstrated that the addition of the above mentioned three components to the fermentation medium could significantly stimulate the biosynthesis of vitamin B₁₂. The concentrations of zinc sulphate, cobaltous chloride and DMBI in the fermentation medium were further optimized with rotatable orthogonal central composite design and statistical analysis by Data Processing System (DPS) software. As a result, vitamin B₁₂ production was increased from 69.36 ± 0.66 to 78.23 ± 0.92 μg/ml.     

Performance of a generalist grasshopper on a C<Subscript>3</Subscript> and a C<Subscript>4</Subscript> grass: compensation for the effects of elevated CO<Subscript>2</Subscript> on plant nutritional quality

The increasing CO₂ concentration in Earths atmosphere is expected to cause a greater decline in the nutritional quality of C₃ than C₄ plants. As a compensatory response, herbivorous insects may increase their feeding disproportionately on C₃ plants. These hypotheses were tested by growing the grasses Lolium multiflorum C₃) and Bouteloua curtipendula C₄) at ambient (370 ppm) and elevated (740 ppm) CO₂ levels in open top chambers in the field, and comparing the growth and digestive efficiencies of the generalist grasshopper Melanoplus sanguinipes on each of the four plant × CO₂ treatment combinations. As expected, the nutritional quality of the C₃ grass declined to a greater extent than did that of the C₄ grass at elevated CO₂; protein levels declined in the C₃ grass, while levels of carbohydrates (sugar, fructan and starch) increased. However, M. sanguinipes did not significantly increase its consumption rate to compensate for the lower nutritional quality of the C₃ grass grown under elevated CO₂. Instead, these grasshoppers appear to use post-ingestive mechanisms to maintain their growth rates on the C₃ grass under elevated CO₂. Consumption rates of the C₃ and C₄ grasses were also similar, demonstrating a lack of compensatory feeding on the C₄ grass. We also examined the relative efficiencies of nutrient utilization from a C₃ and C₄ grass by M. sanguinipes to test the basis for the C₄ plant avoidance hypothesis. Contrary to this hypothesis, neither protein nor sugar was digested with a lower efficiency from the C₄ grass than from the C₃ grass. A novel finding of this study is that fructan, a potentially large carbohydrate source in C₃ grasses, is utilized by grasshoppers. Based on the higher nutrient levels in the C₃ grass and the better growth performance of M. sanguinipes on this grass at both CO₂ levels, we conclude that C₃ grasses are likely to remain better host plants than C₄ grasses in future CO₂ conditions.     

Concentration-Dependent Effects on Intracellular and Surface pH of Exposing <Emphasis Type="Italic">Xenopus</Emphasis> oocytes to Solutions Containing NH<Subscript>3</Subscript>/NH<Subscript>4</Subscript><Superscript>+</Superscript>

Others have shown that exposing oocytes to high levels of (10–20 mM) causes a paradoxical fall in intracellular pH (pH_i), whereas low levels (e.g., 0.5 mM) cause little pH_i change. Here we monitored pH_i and extracellular surface pH (pH_S) while exposing oocytes to 5 or 0.5 mM NH₃/NH₄ ⁺. We confirm that 5 mM causes a paradoxical pH_i fall (−ΔpH_i ≅ 0.2), but also observe an abrupt pH_S fall (−ΔpH_S ≅ 0.2)—indicative of NH₃ influx—followed by a slow decay. Reducing [NH₃/NH₄ ⁺] to 0.5 mM minimizes pH_i changes but maintains pH_S changes at a reduced magnitude. Expressing AmtB (bacterial Rh homologue) exaggerates −ΔpH_S at both levels. During removal of 0.5 or 5 mM NH₃/NH₄ ⁺, failure of pH_S to markedly overshoot bulk extracellular pH implies little NH₃ efflux and, thus, little free cytosolic NH₃/NH₄ ⁺. A new analysis of the effects of NH₃ vs. NH₄ ⁺ fluxes on pH_S and pH_i indicates that (a) NH₃ rather than NH₄ ⁺ fluxes dominate pH_i and pH_S changes and (b) oocytes dispose of most incoming NH₃. NMR studies of oocytes exposed to ¹⁵N-labeled show no significant formation of glutamine but substantial accumulation in what is likely an acid intracellular compartment. In conclusion, parallel measurements of pH_i and pH_S demonstrate that NH₃ flows across the plasma membrane and provide new insights into how a protein molecule in the plasma membrane—AmtB—enhances the flux of a gas across a biological membrane.

Inhibitory role of TGIF in the As<Subscript>2</Subscript>O<Subscript>3</Subscript>-regulated p21<Superscript><Emphasis Type="Italic">WAF1/CIP1</Emphasis></Superscript> expression

Although arsenic is an infamous carcinogen, it has been effectively used to treat acute promyelocytic leukemia, and can induce cell cycle arrest or apoptosis in human solid tumors. Previously, we had demonstrated that opposing effects of ERK1/2 and JNK on p21 expression in response to arsenic trioxide (As₂O₃) are mediated through the Sp1 responsive elements of the p21 promoter in A431 cells. Presently, we demonstrate that Sp1, and c-Jun functionally cooperate to activate p21 promoter expression through Sp1 binding sites (−84/−64) by using DNA affinity binding, chromatin immunoprecipitation, and promoter assays. Surprisingly, As₂O₃-induced c-Jun(Ser63/73) phosphorylation can recruit TGIF/HDAC1 to the Sp1 binding sites and then suppress p21 promoter activation. We suggest that, after As₂O₃ treatment, the N-terminal domain of c-Jun phosphorylation by JNK recruits TGIF/HDAC1 to the Sp1 sites and then represses p21 expression. That is, TGIF is involved in As₂O₃-inhibited p21 expression, and then blocks the cell cycle arrest.     

Cross-talks between Ca<Superscript>2+</Superscript>/CaM and H<Subscript>2</Subscript>O<Subscript>2</Subscript> in abscisic acid-induced antioxidant defense in leaves of maize plants exposed to water stress

Here we examined whether Ca²⁺/Calmodulin (CaM) is involved in abscisic acid (ABA)-induced antioxidant defense and the possible relationship between CaM and H₂O₂ in ABA signaling in leaves of maize (Zea mays L.) plants exposed to water stress. An ABA-deficient mutant vp5 and its wild type were used for the experimentation. We found that water stress enhanced significantly the contents of CaM and H₂O₂, and the activities of chloroplastic and cytosolic superoxide dismutase (SOD), ascorbate peroxidase (APX) and glutathione reductase (GR), and the gene expressions of the CaM1, cAPX, GR1 and SOD4 in leaves of wild-type maize. However, the increases mentioned above were almost arrested in vp5 plants and in the wild-type plants pretreated with ABA biosynthesis inhibitor tungstate (T), suggesting that ABA is required for water stress-induced H₂O₂ production, the enhancement of CaM content and antioxidant defense. Besides, we showed that the up-regulation of water stress-induced antioxidant defense was almost completely blocked by pretreatment with Ca²⁺ inhibitors, CaM antagonists and reactive oxygen (ROS) manipulators. Moreover, the analysis of time course of CaM and H₂O₂ production under water stress showed that the increase in CaM content preceded that of H₂O₂. These results suggested that Ca²⁺/CaM and H₂O₂ were involved in the ABA-induced antioxidant defense under water stress, and the increases of Ca²⁺/CaM contents triggered H₂O₂ production, which inversely affected the contents of CaM. Thus, a cross-talk between Ca²⁺/CaM and H₂O₂ may play a pivotal role in the ABA signaling.     

Pre-treatment with H<Subscript>2</Subscript>O<Subscript>2</Subscript> induces salt tolerance in Barley seedlings

Barley seedlings were pre-treated with 1 and 5 μM H₂O₂ for 2 d and then supplied with water or 150 mM NaCl for 4 and 7 d. Exogenous H₂O₂ alone had no effect on the proline, malondialdehyde (MDA) and H₂O₂ contents, decreased catalase (CAT) activity and had no effect on peroxidase (POX) activity. Three new superoxide dismutase (SOD) isoenzymes appeared in the leaves as a result of 1 μM H₂O₂ treatment. NaCl enhanced CAT and POX activity. SOD activity and isoenzyme patterns were changed due to H₂O₂ pre-treatment, NaCl stress and leaf ageing. In pre-treated seedlings the rate of ¹⁴CO₂ fixation was higher and MDA, H₂O₂ and proline contents were lower in comparison to the seedlings subjected directly to NaCl stress. Cl⁻ content in the leaves 4 and 7 d after NaCl supply increased considerably, but less in pre-treated plants. It was suggested that H₂O₂ metabolism is involved as a signal in the processes of barley salt tolerance.     

Purification and characterization of a cytosolic Ca<Superscript>2+</Superscript>-independent phospholipase A<Subscript>2</Subscript> from bovine brain

The Ca²⁺-independent phospholipase A₂ (iPLA₂) subfamily of enzymes is associated with arachidonic acid (AA) release and the subsequent increase in fatty acid turnover. This phenomenon occurs not only during apoptosis but also during inflammation and lymphocyte proliferation. In this study, we purified and characterized a novel type of iPLA₂ from bovine brain. iPLA₂ was purified 4,174-fold from the bovine brain by a sequential process involving DEAE-cellulose anion exchange, phenyl-5PW hydrophobic interaction, heparin-Sepharose affinity, Sephacryl S-300 gel filtration, Mono S cation exchange, Mono Q anion exchange, and Superose 12 gel filtration. A single peak of iPLA₂ activity was eluted at an apparent molecular mass of 155 kDa during the final Superose 12 gel-filtration step. The purified enzyme had an isoelectric point of 5.3 on twodimensional gel electrophoresis (2-DE) and was inhibited by arachidonyl trifluoromethyl ketone (AACOCF₃), Triton X-100, iron, and Ca²⁺. However, it was not inhibited by bromoenol lactone (BEL), an inhibitor of iPLA₂, and adenosine triphosphate (ATP). The spot with the iPLA₂ activity did not match with any known protein sequence, as determined by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) analysis. Altogether, these data suggest that the purified enzyme is a novel form of cytosolic iPLA₂.     

Cellular expression of C<Subscript>3</Subscript> and C<Subscript>4</Subscript> photosynthetic enzymes in the amphibious sedge<Emphasis Type="Italic"> Eleocharis retroflexa</Emphasis> ssp.<Emphasis Type="Italic"> chaetaria</Emphasis>

The amphibious leafless sedge Eleocharis retroflexa ssp. chaetaria expresses C₄-like biochemical characteristics in both the terrestrial and submerged forms. Culms of the terrestrial form have Kranz anatomy, whereas those of the submerged form have Kranz-like anatomy combined with anatomical features of aquatic plant leaves. We examined the immunolocalization of C₃ and C₄ enzymes in culms of the two forms. In both forms, phosphoenolpyruvate carboxylase; pyruvate, Pi dikinase; and NAD-malic enzyme were compartmentalized between the mesophyll (M) and Kranz cells, but their levels were somewhat reduced in the submerged form. In the terrestrial form, ribulose-1,5-bisphosphate carboxylase/oxygenase (rubisco) occurred mainly in the Kranz cells, and weakly in the M chloroplasts. In the submerged form, the rubisco occurred at higher levels in the M cells than in the terrestrial form. In both forms, the C₄ pattern of enzyme expression was clearer in the M cells adjacent to Kranz cells than in distant M cells. During the transition from terrestrial to submerged conditions, the enzyme expression pattern changed in submerged mature culms that had been formed in air before submergence, and matched that in culms newly developed underwater. It seems that effects of both environmental and developmental factors overlap in the C₄ pattern expression in this plant.     

Enhancing yield of <Emphasis Type="Italic">S</Emphasis>-adenosylmethionine in <Emphasis Type="Italic">Pichia pastoris</Emphasis> by controlling NH<Subscript>4</Subscript><Superscript>+</Superscript> concentration

Yield of S-adenosylmethionine was improved significantly in recombinant Pichia pastoris by controlling NH₄ ⁺ concentration. The highest production rate was 0.248 g/L h when NH₄ ⁺ concentration was 450 mmol/L and no repression of cell growth was observed. Within very short induction time (47 h), 11.63 g/L SAM was obtained in a 3.7 L bioreactor.     

Ca<Superscript>2+</Superscript> increases the specific coenzyme Q<Subscript>10</Subscript> content in <Emphasis Type="Italic">Agrobacterium tumefaciens</Emphasis>

Of various metal ions (Ca²⁺, Cr³⁺, Cu²⁺, Fe²⁺, Mg²⁺, Mn²⁺, Ni²⁺ and Zn²⁺) added to the culture medium of Agrobacterium tumefaciens at 1 mM, only Ca²⁺ increased Coenzyme Q10 (CoQ₁₀) content in cells without the inhibition of cell growth. In a pH-stat fed-batch culture, supplementation with 40 mM of CaCO₃ increased the specific CoQ₁₀ content and oxidative stress by 22.4 and 48%, respectively. Also, the effect of Ca²⁺ on the increase of CoQ₁₀ content was successfully verified in a pilot-scale (300 L) fermentor. In this study, the increased oxidative stress in A. tumefaciens culture by the supplementation of Ca²⁺ is hypothesized to stimulate the increase of specific CoQ₁₀ content in order to protect the membrane against lipid peroxidation. Our results improve the understanding of Ca²⁺ effect on CoQ₁₀ biosynthesis in A. tumefaciens and should contribute to better industrial production of CoQ₁₀ by biological processes.     

Ginsenoside Rg<Subscript>3</Subscript> enhances large conductance Ca<Superscript>2+</Superscript>-activated potassium channel currents: A role of Tyr360 residue

Ginsenosides, active ingredients of Panax ginseng, are known to exhibit neuroprotective effects. Large-conductance Ca²⁺-activated K⁺ (BK_Ca) channels are key modulators of cellular excitability of neurons and vascular smooth muscle cells. In the present study, we examined the effects of ginsenosides on rat brain BK_Ca (rSlo) channel activity heterologously expressed in Xenopus oocytes to elucidate the molecular mechanisms how ginsenoside regulates the BK_Ca channel activity. Ginsenoside Rg₃ (Rg₃) enhanced outward BK_Ca channel currents. The Rg₃-enhancement of outward BK_Ca channel currents was concentration-dependent, voltage-dependent, and reversible. The EC₅₀ was 15.1 ± 3.1 μM. Rg₃ actions were not desensitized by repeated treatment. Tetraetylammonium (TEA), a K⁺ channel blocker, inhibited BK_Ca channel currents. We examined whether extracellular TEA treatment could alter the Rg₃ action and vice versa. TEA caused a rightward shift of the Rg₃ concentration-response curve (i.e., much higher concentration of Rg₃ is required for the activation of BK_Ca channel compared to the absence of TEA), while Rg₃ caused a rightward shift of the TEA concentration-response curve in wild-type channels. Mutation of the extracellular TEA binding site Y360 to Y360I caused a rightward shift of the TEA concentration-response curve and almost abolished both the Rg₃ action and Rg₃-induced rightward shift of TEA concentration-response curve. These results indicate that Tyr360 residue of BK_Ca channel plays an important role in the Rg₃-enhancement of BK_Ca channel currents.     

Vitamin B<Subscript>12</Subscript>-independent strains of <Emphasis Type="Italic">Methylophaga marina</Emphasis> isolated from Red Sea algae

Two strains (KM3 and KM5) of halophilic methylobacteria isolated from Red Sea algae do not require vitamin B₁₂ for growth and can use methanol, methylamine, dimethylamine, trimethylamine, dimethyl sulfide, and fructose as sources of carbon and energy. The cells of these strains are gram-negative motile monotrichous (strain KM3) or peritrichous (strain KM5) rods. The strains are strictly aerobic and require Na⁺ ions but not growth factors. They are oxidase-and catalase-positive and reduce nitrates to nitrites. Both strains can grow in a temperature range of 4 to 37°C (with optimal growth at 29–34°C), at pH between 5.5 and 8.5 (with optimal growth at pH 7.5–8.0), and in a range of salt concentrations between 0.5 and 15% NaCl (with optimal growth at 5–9% NaCl). The phospholipids of these strains are dominated by phosphatidylethanolamine and phosphatidylglycerol and also include phosphatidylcholine, phosphatidylserine, and cardiolipin. The dominant fatty acids are C_16:1ω7c and C_16:0. The major ubiquinone is Q₈. The cells accumulate ectoin, glutamate, and sucrose as intracellular osmoprotectants. The strains implement the 2-keto-3-deoxy-6-phosphogluconate-dependent variant of the ribulose monophosphate pathway. The G+C content of the DNA is 44.4–44.7 mol%. Analysis of the 16S rRNA genes showed that both strains belong to Gammaproteobacteria and have a high degree of homology (99.4%) to Methylophaga marina ATCC 35842^T. Based on the data of polyphasic taxonomy, isolates KM3 and KM5 are identified as new strains M. marina KM3 (VKM B-2386) and M. marina KM5 (VKM B-2387). The ability of these strains to produce auxins (indole-3-acetic acid) suggests their metabolic association with marine algae.     

Fumonisin B<Subscript>1</Subscript>, a sphingoid toxin,is a potent inhibitor of the plasma membrane H<Superscript>+</Superscript>-ATPase

Fumonisin B₁ (FB₁) is an amphipathic toxin produced by the pathogenic fungus Fusarium verticillioides which causes stem, root and ear rot in maize (Zea mays L.). In this work, we studied the action of FB₁ on the plasma membrane H⁺-ATPase (EC 3.6.1.34) from germinating maize embryos, and on the fluidity and lipid peroxidation of these membranes. In maize embryos the toxin at 40 M inhibited root elongation by 50% and at 30 M decreased medium acidification by about 80%. Irrespective of the presence and absence of FB₁, the H⁺-ATPase in plasma membrane vesicles exhibited non-hyperbolic saturation kinetics by ATPH-Mg, with Hill number of 0.67. Initial velocity studies revealed that FB₁ is a total uncompetitive inhibitor of this enzyme with an inhibition constant value of 17.5±1 M. Thus FB₁ decreased V_max and increased the apparent affinity of the enzyme for ATP-Mg to the same extent. Although FB₁ increased the fluidity at the hydrophobic region of the membrane, no correlation was found with its effect on enzyme activity, since both effects showed different FB₁-concentration dependence. Peroxidation of membrane lipids was not affected by the toxin. Our results suggest that, under in vivo conditions, the plasma membrane H⁺-ATPase is a potentially important target of the toxin, as it is inhibited not only by FB₁ but also by its structural analogs, the sphingoid intermediates, which accumulate upon the inhibition of sphinganine N-acyltransferase by this toxin.     

Genomics of the<Emphasis Type="Italic"> ccoNOQP</Emphasis>-encoded<Emphasis Type="Italic"> cbb</Emphasis><Subscript>3</Subscript> oxidase complex in bacteria

Many bacteria adapt to microoxic conditions by synthesizing a particular cytochrome c oxidase (cbb ₃) complex with a high affinity for O₂, encoded by the ccoNOQP operon. A survey of genome databases indicates that ccoNOQP sequences are widespread in all sub-branches of Proteobacteria but otherwise are found only in bacteria of the CFB group (Cytophaga, Flexibacter, Bacteroides). Our analysis of available genome sequences suggests four major strategies of regulating ccoNOQP expression in response to O₂. The most widespread strategy involves direct regulation by the O₂-responsive protein Fnr. The second strategy involves an O₂-insensitive paralogue of Fnr, FixK, whose expression is regulated by the O₂-responding FixLJ two-component system. A third strategy of mixed regulation operates in bacteria carrying both fnr and fixLJ-fixK genes. Another, not yet identified, strategy is likely to operate in the -Proteobacteria Helicobacter pylori and Campylobacter jejuni which lack fnr and fixLJ-fixK genes. The FixLJ strategy appears specific for the -subclass of Proteobacteria but is not restricted to rhizobia in which it was originally discovered.     

Contribution of the [Fe<Superscript>II</Superscript>(SCys)<Subscript>4</Subscript>] site to the thermostability of rubredoxins

The thermostabilities of Fe²⁺ ligation in rubredoxins (Rds) from the hyperthermophile Pyrococcus furiosus (Pf) and the mesophiles Clostridium pasteurianum (Cp) and Desulfovibrio vulgaris (Dv) were compared. Residue 44 forms an NH...S(Cys) hydrogen bond to one of the cysteine ligands to the [Fe(SCys)₄] site, and substitutions at this location affect the redox properties of the [Fe(SCys)₄] site. Both Pf Rd and Dv Rd have an alanine residue at position 44, whereas Cp Fd has a valine residue. Wild-type proteins were examined along with V44A and A44V exchange mutants of Cp and Pf Rds, respectively, in order to assess the effects of the residue at position 44 on the stability of the [Fe(SCys)₄] site. Stability of iron ligation was measured by temperature-ramp and fixed-temperature time course experiments, monitoring iron release in both the absence and presence of more thiophilic metals (Zn²⁺, Cd²⁺) and over a range of pH values. The thermostability of the polypeptide fold was concomitantly measured by fluorescence, circular dichroism, and ¹H NMR spectroscopies. The A44V mutation strongly lowered the stability of the [Fe^II(SCys)₄] site in Pf Rd, whereas the converse V44A mutation of Cp Rd significantly raised the stability of the [Fe^II(SCys)₄] site, but not to the levels measured for wild-type Dv Rd. The region around residue 44 is thus a significant contributor to stability of iron coordination in reduced Rds. This region, however, made only a minor contribution to the thermostability of the protein folding, which was found to be higher for hyperthermophilic versus mesophilic Rds, and largely independent of the residue at position 44. These results, together with our previous studies, show that localized charge density, solvent accessibility, and iron site/backbone interactions control the thermostability of the [Fe(SCys)₄] site. The iron site thermostability does make a minor contribution to the overall Rd thermostability. From a mechanistic standpoint, we also found that attack of displacing ions (H⁺, Cd²⁺) on the Cys42 sulfur ligand at the [Fe(SCys)₄] site occurs through the V8 side and not the V44 side of the iron site.Electronic Supplementary Material Supplementary material is available in the online version of this article at http://dx.doi.org/10.1007/s00775-004-0525-4Abbreviations BPS bathophenanthroline sulfonate, sodium salt - Cp Rd (Pf Rd, Dv Rd) recombinant rubredoxin from Clostridium pasteurianum (Pyrococcus furiosus, Desulfovibrio vulgaris) - HEPES hydroxyethylpiperazineethanesulfonic acid - MES morpholinoethanesulfonic acid - Tris tris(hydroxymethyl)aminomethane - wt wild-type - ZnRd recombinant rubredoxin containing a [Zn(SCys)₄] site     

<Emphasis Type="Italic">Undaria pinnatifida</Emphasis> fucoidan extract protects against CCl<Subscript>4</Subscript>-induced oxidative stress

This study was performed to elucidate the effects of Undaria pinnatifida fucoidan extract (UPFE) in preventing CCl₄-induced oxidative stress. UPFE (100 mg/kg) was intraperitoneally administered to rats for 14 days. On day 15, CCl₄ dissolved in olive oil (50% CCl₄) was injected 12 h before they were anesthetized and dissected. To measure UPFE-mediated antioxidation, we examined the levels of glutamate oxaloacetate transaminase (GOT), glutamate pyruvate transaminase (GPT), alkaline phosphatase (ALP), and lactate dehydrogenase (LDH) in serum, as well as malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) in liver homogenates. CCl₄ treatment markedly increased the levels of GOT, GPT, ALP, LDH, and MDA and significantly decreased levels of SOD, CAT, and GPx. UPFE pretreatment decreased levels of GOT, GPT, ALP, LDH, and MDA, by 62.8, 68.5, 41.9, 72.7, and 122%, respectively and increased those of SOD, CAT, and GPx by 111.1, 15.9, and 52.6%, respectively. These results showed that UPFE has antioxidant effects against CCl₄-induced oxidative stress.     

Mitochondrial and cell-surface F<Subscript>0</Subscript>F<Subscript>1</Subscript>ATPsynthase in innate and acquired cardioprotection

Mitochondria are central to heart function and dysfunction, and the pathways activated by different cardioprotective interventions mostly converge on mitochondria. In a context of perspectives in innate and acquired cardioprotection, we review some recent advances in F₀F₁ATPsynthase structure/function and regulation in cardiac cells. We focus on three topics regarding the mitochondrial F₀F₁ATPsynthase and the plasma membrane enzyme, i.e.: i) the crucial role of cardiac mitochondrial F₀F₁ATPsynthase regulation by the inhibitory protein IF₁ in heart preconditioning strategies; ii) the structure and function of mitochondrial F₀F₁ATPsynthase oligomers in mammalian myocardium as possible endogenous factors of mitochondria resistance to ischemic insult; iii) the external location and characterization of plasma membrane F₀F₁ ATP synthase in search for possible actors of its regulation, such as IF₁ and calmodulin, at cell surface.