The measurement of intracellular sodium activities in the bullfrog by means of double-barreled sodium liquid ion-exchanger microelectrodes.

A double-barreled Na+-selective microelectrode was constructed with monensin as a liquid ion exchanger. The HCl-treated monensin was dissolved in a solvent (Corning 477317) at 10% (weight/weight). Internal reference solution of its ionic barrel was mixture of 0.49 M NaCl and 0.01 M KCl, the pH being adjusted to 3 with 0.1 M citrate-HCl buffer, whereas that of the PD barrel was 0.5 M KCl. Average slope and selectivity ratio (Na+/K+) tested on 10 different microelectrodes were -57.5 +/- 1.87 mV/P(Na) (SEM) and 6.7 +/- 0.44, respectively. The electrical resistance was an order of 10(10) ohm and the response time was less than 10 sec. Using this microelectrode, a free flow micropuncture experiment was carried out in the bullfrog kidney and the intracellular Na+ activity as well as the membrane PD was determined on the proximal tubular cell. Average value (+/- SEM, n = 15) for the intracellular Na+ and K+ was 20.7 +/- 1.56 mEq/L and 61.2 +/- 1.16 mEq/L, respectively, and -68.7 +/- 0.88 mV for the peritubular membrane PD. There was a significant negative correlation between Na+ and K+ activities within the cell, i.e., the lower the ionic activity of cellular Na+ was, the higher the cellular K+, and vice versa, the sum of these two being kept nearly constant. The above finding may be somehow related to the isosmosis in the reabsorptive process across the proximal tubular epithelium.     

Studies of oligosaccharide linkages by simultaneous determination of component aldehydes in dialdehyde fragments as (2,4-dinitrophenyl)hydrazones

The component aldehydes in dialdehyde fragments formed by periodate oxidation of oligosaccharides were converted quantitatively into the corresponding (2,4-dinitrophenyl)hydrazones by the simple procedure of treatment with excess (2,4-dinitrophenyl)hydrazine hydrochloride in 1,2-dimethoxyethane. Then, by chromatographic separation of the hydrazones on a small column of silica gel and subsequent spectrophotometric analysis, it was possible to determine the position of glycosidic substitution in μmolar amounts of various types of glucobioses, oligosaccharides of senega, and some synthetic (1→6)-β-D-gluco-oligosaccharides.     

Sites of in vivo histone methylation in developing trout testis.

Specific lysyl residues of trout testis histones H3 and H4 are methylated partially during rainbow trout spermatogenesis. Histones H1, H2A, H2B, and protamine are not methylated. The single site (lysine 20) in histone H4 and the two major sites (lysines 9 and 27) in histone H3 are homologous to those determined for other organisms, but an additional minor site (lysine 4) occurs in histone H3. As described for calf thymus, both histones H3 and H4 contain epsilon-N-mono- and dimethyllysine, while histone H3 contains in addition, epsilon-N-trimethyllysine. The trout-specific histone H6, which accounts for 0.5 to 1.0% of total histone, contains a sequence for residues 3 to 5,-Arg-Lys-Ser-, which is the same as one methylated in histones H3, at lysines 9 and 27. However, histone H6 yields only trace amounts of [3H]methyl incorporation and no detectable methyllysines on amino acid analysis.     

Properties of chromatin subunits from developing trout testis.

When a sample of trout testis nuclei is digested with micrococcal nuclease, the DNA is cleaved almost entirely to discrete fragments approximately 200 base pairs long and multiples thereof. The same DNA fragments can be obtained when isolated chromatin, as opposed to intact nuclei, is nuclease digested. These DNA fragments can also be found in discrete chromatin "subunits" isolated from nuclease-digested nuclei. Sedimentation through sucrose gradients or velocity sedimentation in an analytical ultracentrifuge separates these chromatin subunits into 11 S (monomer), 16 S (dimer), and 22 S (trimer) etc. species. Subunits can also be fractionated on a Sepharose 2B column equilibrated and run in low salt. High salt (greater than 40 mM NaCl) or divalent cations (congruent to 5 mM) cause subunit precipitation. Chromatin subunits have a protein to DNA ratio of approximately 1.2 and contain all the histones, including the trout-specific histone T. There are, however, no detectable nonhistone chromosomal proteins. Mg-2+ precipitates of the 11 S chromatin monomers, when pelleted, are thin and clear, while oligomer Mg-2+ pellets are thick and white. This could reflect a more symmetrical or ordered packing of 11 S monomers, which are deficient in histone I. This histone may cross-link the larger oligomers, resulting in a disordered Mg-2+ complex. These results are consistent with the subunit model of chromatin structure, based on 200 base pair long regions of DNA associated with histones. These subunits would be separated by nuclease-sensitive DNA spacer regions and cross-linked by histone I.     

Proteoglycan complexes from bovine heart valve. Fractionation by density-gradient centrifugation and gel filtration under dissociative conditions.

Proteoglycan complexes from collagenase [EC 3.4.24.3]-indigestible materials of bovine heart valves were extracted with 4 M guanidinium chloride, purified by ion-exchange column chromatography in a urea-containing solution, then fractionated by density-gradient centrifugation under dissociative conditions. Electrophoretic characteristics and enzymic susceptibility of the density-gradient fractions revealed that the glycosaminoglycans constituting the proteoglycan complexes in this indigestible materials were mainly dermatan sulfate in the top three fractions, and dermatan sulfate and chondroitin sulfates in the bottom fraction; a minor constituent which was common to all the fractions was hyaluronic acid. A gel-like substance (Fr. Ig) at the top of the gradient, amounting to about 25% of the loaded dry sample, contained only a trace of hydroxyproline (less than 1%) and was composed of proteodermatan sulfate, glycoprotein, and a small amount of hyaluronic acid. Sodium dodecyl sulfate-polyacrylamide gel electrophoretic analyses of Fr. Ig with 2-mercaptoethanol showed that the major part of the proteins in this gel-like substance was cross-linked by disulfide bridges. Chromatography of Fr. Ig on Sepharose 4B in buffered 4 M guanidinium chloride containing 2-mercaptoethanol, together with the electrophoretic patterns of the resulting fractions, suggested that proteodermatan sulfate was not associated with hyaluronic acid through covalent bonds. The amino acid composition of Fr. Ig was very similar to that reported in the literature for "dermatan sulfate-protein complex", and "structural glycoprotein" or "acidic structural protein".     

Production of Antibacterial Compounds Analogous to Chloramphenicol by a n-Paraffin-grown Bacterium

Corynebacterium sp. KY 4339, when grown on n-paraffin (a mixture of C–12 to C–14 fractions) as the sole carbon source, produced three kinds of antibacterial compounds which were tentatively named Corynecins. These compounds were isolated by the extraction from the culture broth with ethyl acetate and by the chromatographies on silicic acid and alumina columns. Each component demonstrated some similarity to chloramphenicol on thin-layer chromatogram. Although their biological activities were not so remarkably as that of chloramphenicol, the patterns of antibacterial spectra against gram-positive and gram-negative bacteria resembled to it.

For the production of corynecins, n-paraffin was a preferable carbon source. By controlling the pH of the medium in the neutral range and keeping the aeration at a high level during the fermentation, approximately 3 g of corynecins per liter of the medium were produced after 72-hr incubation.     

Organelle contacts: Sub‐organelle zones to facilitate rapid and accurate inter‐organelle trafficking of lipids

When one person wants to communicate securely with another, he/she should contact the other person directly. This rule applies not only to human society, but also to the intracellular micro‐society. In the past two decades, it has become increasingly clear that the sub‐organelle regions called membrane contact sites (MCSs) are pivotal for inter‐organelle transport of lipids in cells, as highlighted in the thematic review series “Interorganelle trafficking of lipids” held in Traffic in 2014–2015. In this commentary, we will describe how the currently prevailing model for lipid trafficking at MCSs was generated, and comment on three important issues that have not been explored: (a1) the principles guiding the generation of an asymmetrical inter‐organelle flow of lipids in cells, (b2) the advantages in lipid trafficking at organelle contacts, and (c3) the dynamic network of inter‐organelle lipid trafficking.     

Focus: Zoonotic Disease: New Series of Imidazoles Showed Promising Growth Inhibitory and Curative Potential Against Trypanosoma Infection

The Trypanosoma spp. cause animal and human trypanosomiasis characterized with appreciable health and economic burden mostly in developing nations. There is currently no effective therapy for this parasitic disease, due to poor drug efficacy, drug resistance, and unwanted toxicity, etc. Therefore, new anti-Trypanosoma agents are urgently needed. This study explored new series of imidazoles for anti-Trypanosoma properties in vitro and in vivo. The imidazoles showed moderate to strong and specific action against growth of T. congolense. For example, the efficacy of the imidazole compounds to restrict Trypanosoma growth in vitro was ≥ 12-fold specific towards T. congolense relative to the mammalian cells. Additionally, the in vivo study revealed that the imidazoles exhibited promising anti-Trypanosoma efficacy corroborating the in vitro anti-parasite capacity. In particular, three imidazole compounds (C1, C6, and C8) not only cleared the systemic parasite burden but cured infected rats after no death was recorded. On the other hand, the remaining five imidazole compounds (C2, C3, C4, C5, and C7) drastically reduced the systemic parasite load while extending survival time of the infected rats by 14 days as compared with control. Untreated control died 3 days post-infection, while the rats treated with diminazene aceturate were cured comparable to the results obtained for C1, C6, and C8. In conclusion, this is the first study demonstrating the potential of these new series of imidazoles to clear the systemic parasite burden in infected rats. Furthermore, a high selectivity index of imidazoles towards T. congolense in vitro and the oral LD₅₀ in rats support anti-parasite specific action. Together, findings support the anti-parasitic prospects of the new series of imidazole derivatives.