The role of the medium in long-range electron transfer

 The role of the polypeptide matrix in electron transfer processes in proteins has been studied in two distinct systems: first in a protein where the induced ET is artificial, and second as part of the catalytic cycle of an enzyme. Azurins are structurally well-characterized blue single-copper proteins consisting of a rigid β-sheet polypeptide matrix. We have determined rate constants and activation parameters for intramolecular long-range electron transfer between the disulfide radical anions (generated by pulse radiolysis) and the copper(II) centre as a function of driving force and nature of the intervening medium in a large number of wild-type and single-site-mutated proteins. In ascorbate oxidase, for which the three-dimensional structure is equally well characterized, the internal ET from the type-I Cu(I) to the trinuclear Cu(II) centre has been studied. We find that the results correlate well with distance through well-defined pathways using a through-bond electron tunnelling mechanism. Received: 2 January 1997 / Accepted: 6 February 1997     

Mitochondrial DNA haplogrouping of the brown bear,Ursus arctos (Carnivora: Ursidae) in Asia,based on a newly developed APLP analysis

Sequence analyses of the complete brown bear, Ursus arctos, mitochondrial DNA (mtDNA) genome have detected scattered single nucleotide polymorphisms (SNPs) that define distinct mtDNA haplogroups in phylogeographical studies. The degraded DNA in historical samples, such as stuffed or excavated specimens, however, is often not suitable for sequence analyses. To address this problem, we developed an amplified product length polymorphism (APLP) analysis for mtDNA‐haplogrouping U. arctos specimens by detecting haplogroup‐specific SNPs. We verified the validity and utility of this method by analysing up to 170‐year‐old skin samples from U. arctos specimens collected widely across continental Eurasia. We detected some of the same haplogroups as those occurring in eastern Hokkaido (Japan) and eastern Alaska in continental Eurasia (the Altai and the Caucasus). Our results show that U. arctos in eastern Hokkaido and eastern Alaska descended from a common ancestor in continental Eurasia, and suggest that U. arctos occupied several refugia in southern Asia during the Last Glacial Maximum. © 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 111 , 627–635.     

Comparison of changes in succinate dehydrogenase activity in blood lymphocytes and modification of radiosensitivity by exogenous hypoxia

Radioprotective efficiency of gas hypoxic mixtures (GHM) containing 5-12% of oxygen and the rate of the reaction of succinate dehydrogenase (VSDG) activity in peripheral blood lymphocytes upon breathing GHM were comparatively studied in rats and dogs. VSDG was 4393.5 (%O2)-2.58 and 130.76 (%O2)-1.42 in dogs and rats respectively. Taking into account that DMF in rats is a function of oxygen concentration in the mixture one can obtain a formula for determining a dose modifying factor (DMF) as a function of the rate of SDG activity reaction.     

Calcium-Induced Desensitization of Acetylcholine Release from Synaptosomes or Proteoliposomes Equipped with Mediatophore, a Presynaptic Membrane Protein

A "fatigue" of acetylcholine (ACh) release is described in cholinergic synaptosomes stimulated with the calcium ionophore A23187 or gramicidin. A small conditioning calcium entry, which did not trigger a large ACh release, led to a decrease of transmitter release elicited by a second large calcium influx. This fatigue was half-maximal at approximately 30 microM external calcium and developed in a few minutes. In contrast, activation of release by calcium was very rapid and was half-maximal at approximately 0.5 mM external calcium. Activation and desensitization of release could be attributed to the recently identified presynaptic membrane protein, the "mediatophore." Proteoliposomes equipped with purified mediatophore showed a calcium-dependent activation and "fatigue" of ACh release similar to that of synaptosomes. It was found that the ionophore A23187 rapidly equilibrated internal and external calcium concentrations in proteoliposomes. Thus, the external calcium concentration gave the internal concentration required for activation or desensitization of proteoliposomal ACh release. The mediatophore showed remarkable calcium binding properties (20 sites/molecule) with a KD of 25 microM. The physiological implications of desensitization on the organization of release sites are discussed.     

Characterization by cytofluorometry of the different types of adenohypophyseal cells in the rat

The different antehypophysical cell types which synthetize and release somatotroph (GH), corticothroph (ACTH), gonadotroph (LH-FSH) and lactotroph (PRL) hormones were analysed. The experiments were performed on hypophyses from five groups of animals: adult males, 14 days-old female, adult females, gestating adult females and lactating adult females. The cells were analysed by immunofluorescence using flow cytometry. For each of the hormones studied, there was a characteristic spectral distribution of cells. The evolution of cell size and granular content with respect to sex and physiological state of each group was studied by the analysis of diffused light. Small, slightly granular cells represented 50% of the cell population in males and 14 day-old females but only 8% in gestating or lactating females. The study of the cell cycle showed the presence of dividing cells in the population of large, granular cells from gestating and from lactating females. No features of cell division were observed in the population of small, slightly granular cells. This study indicates the potential value of multiparametric analysis in the separation of pure sub-populations of antehypophysial cells.     

Isoenzyme pattern and subcellular localisation of enzymes involved in fumaric acid accumulation by Rhizopus oryzae

Summary Electrophoretic studies of fumarase and nicotine adenine dinucleotide (NAD)-malate dehydrogenase were carried out in the fumaric acid-accumulating fungus Rhizopus oryzae. The analyses revealed two fumarase isoenzymes, one localised solely in the cytosol and the other found both in the cytosol and in the mitochondrial fraction. The activity of the cytosolic isoenzyme of fumarase was higher during the acid production stage than during growth. Addition of cycloheximide inhibited fumaric acid production and decreased the activity of the cytosolic isoenzyme of fumarase. These results suggested that de novo protein synthesis is required for increase in the activity of the cytosolic isoenzyme and that such an increase in activity is essential for fumaric acid accumulation. Three distinct isoenzymes of NAD-malate dehydrogenase could be detected in R. oryzae. No changes were observed in the isoenzyme pattern of malate dehydrogenase during fumaric acid production.