In vitro kinetics of the rat brain succinate dehydrogenase inhibition by hexachlorophene.

Brain succinate dehydrogenase (SDH) activity was inhibited by in vitro hexachlorophene (HCP) with a half inhibitory concentration (IC50) of 0.65 x 10(-3) M. The HCP exerted noncompetitive inhibition at 0.5 mM (IC50) on SDH activity. The brain SDH demands more energy of activation (deltaE) in the presence of HCP. The ionizable groups of SDH such as the sulfhydral group of cysteine and alpha-amino groups of cysteine were not altered qualitatively in the presence of HCP.     

Characterization of synaptosomes from the central nervous system of insects

Nerve terminals from the head ganglia of Locusta migratoria were isolated by means of a modified microscale flotation technique. Enzymatic, ultrastructural and chemical analysis revealed that the synaptosomal fraction was highly enriched in well-preserved nerve endings containing almost no free mitochondria. Cholinergic activities (choline acetyltransferase, acetylcholinesterase, acetylcholine receptors) were found to be concentrated in the synaptosomal fraction. The cholinergic nature and the functional integrity of nerve endings isolated from locusts were further supported by the existence of a high affinity choline uptake system, which is abolished by hemicholinium-3 as well as by low temperature, is essentially sodium dependent and inhibited by elevated potassium concentrations. After slight modifications of the gradient densities, synaptosomes could also be isolated from other insect species.     

Membranes of Rhodopseudomonas sphaeroides. VI. Isolation of a fraction enriched in newly synthesized bacteriochlorophyll a-protein complexes

Radioactivity eventually destined for the chromatophore membrane of Rhodopseudomonas sphaeroides was shown in pulse-chase studies to appear first in a distinct pigmented fraction. This material formed an upper pigmented band which sedimented more slowly than chromatophores when cell-free extracts were subjected directly to rate-zone sedimentation on sucrose density gradients. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that the purified fraction contained polypeptide bands of the same mobility as light-harvesting bacteriochlorophyll a and reaction center-associated protein components of chromatophores; these were superimposed upon cytoplasmic membrane polypeptides. The pulse-chase relation was confined mainly to the polypeptide components of these pigment-protein complexes. It is suggested that the isolated fraction may be derived from sites at which new membrane invagination is initiated.     

Mitochondrial potassium channels and reactive oxygen species

Pretreatment of tissues with potassium channel openers (KCO’s) has been observed to be cytoprotective in a broad variety of insults. This phenomenon has been proposed to be intimately linked to activation of mitochondrial potassium channels which apparently modulate the mitochondrial production of reactive oxygen species (ROS). This critical review summarizes literature findings about the mitochondrial production of ROS, the action of KCO’s on mitochondrial ROS production and the putative link to the cytoprotective action of these drugs.     

Analysis of covalent flavinylation using thermostable succinate dehydrogenase from Thermus thermophilus and Sulfolobus tokodaii lacking SdhE homologs

Recent studies have indicated that post-translational flavinylation of succinate dehydrogenase subunit A (SdhA) in eukaryotes and bacteria require the chaperone-like proteins Sdh5 and SdhE, respectively. How does covalent flavinylation occur in prokaryotes, which lack SdhE homologs? In this study, I showed that covalent flavinylation in two hyperthermophilic bacteria/archaea lacking SdhE, Thermus thermophilus and Sulfolobus tokodaii, requires heat and dicarboxylic acid. These thermophilic bacteria/archaea inhabit hot environments and are said to be genetically far removed from mesophilic bacteria which possess SdhE. Since mesophilic bacteria have been effective at covalent bonding in temperate environments, they may have caused the evolution of SdhE.     

High-lysine corn generated by endosperm-specific suppression of lysine catabolism using RNAi

Because of the limited lysine content in corn grain, synthetic lysine supplements are added to corn meal-based rations for animal feed. The development of biotechnology, combined with the understanding of plant lysine metabolism, provides an alternative solution for increasing corn lysine content through genetic engineering. Here, we report that by suppressing lysine catabolism, transgenic maize kernels accumulated a significant amount of lysine. This was achieved by RNA interference (RNAi) through the endosperm-specific expression of an inverted-repeat (IR) sequence targeting the maize bifunctional lysine degradation enzyme, lysine-ketoglutarate reductase/saccharopine dehydrogenase (ZLKR/SDH). Although plant-short interfering RNA (siRNA) were reported to lack tissue specificity due to systemic spreading, we confirmed that the suppression of ZLKR/SDH in developing transgenic kernels was restricted to endosperm tissue. Furthermore, results from our cloning and sequencing of siRNA suggested the absence of transitive RNAi. These results support the practical use of RNAi for plant genetic engineering to specifically target gene suppression in desired tissues without eliciting systemic spreading and the transitive nature of plant RNAi silencing.     

How Mitochondrial Metabolism Contributes to Macrophage Phenotype and Functions

Metabolic reprogramming of cells from the innate immune system is one of the most noteworthy topics in immunological research nowadays. Upon infection or tissue damage, innate immune cells, such as macrophages, mobilize various immune and metabolic signals to mount a response best suited to eradicate the threat. Current data indicate that both the immune and metabolic responses are closely interconnected. On account of its peculiar position in regulating both of these processes, the mitochondrion has emerged as a critical organelle that orchestrates the coordinated metabolic and immune adaptations in macrophages. Significant effort is now underway to understand how metabolic features of differentiated macrophages regulate their immune specificities with the eventual goal to manipulate cellular metabolism to control immunity. In this review, we highlight some of the recent work that place cellular and mitochondrial metabolism in a central position in the macrophage differentiation program.     

Detection of mitochondrial dysfunction in vitro by laser‐induced autofluorescence

Mitochondrion plays a significant role in a variety of biological functions. Because of their diverse character and location in the cellular systems, mitochondria commonly get exposed to various extrinsic and intrinsic cellular stresses. The present study reports a novel approach to detection of mitochondrial dysfunction based on tryptophan autofluorescence of its proteins in mouse liver, using laser‐induced fluorescence (LIF) as a tool. Mitochondria, isolated from the mouse liver, were initially tested for purity and integrity using lactate dehydrogenase and succinate dehydrogenase (SDH) assays. Mitochondrial stress was induced by treating the isolated mitochondria with heavy metals at 10 and 0.01 mM for sodium arsenite and mercuric chloride, respectively. Upon treatment with the heavy metal, tryptophan autofluorescence quenching was recorded at 281 nm excitation. The functional integrity of the mitochondria treated with heavy metals was evaluated by measuring SDH and cytochrome c oxidase activities at various concentrations of mitochondria, which showed impaired activity as compared to control upto a concentration of 6.25 μg. A significant shift was also observed in the autofluorescence of proteins upto the level below 1 μg, suggesting their conformational change and hence altered structural integrity of mitochondria. Circular dichroism spectroscopy data of the mitochondrial proteins treated with heavy metals further validates their conformational change as compared to untreated control. The present study clearly shows that the LIF can be a novel detection tool to detect altered structural integrity of cellular mitochondria upon stress, and it also possesses the potentiality to combine with other interdisciplinary modalities.