Antibiotic activity of isoxazolylnaphthoquinone imines on mice infected with Staphylococcus aureus

I. ALBESA, P. BOGDANOV, A. ERASO, N.R. SPERANDEO AND M.M. DE BERTORELLO. 1995. The antibiotic activity of new synthetic isoxazolylnaphthoquinone imines was studied. Pseudomonas aeruginosa ATCC 27853 and Escherichia coli ATCC 25922 were resistant to the four compounds studied (MIC > 128 µg ml⁻¹), but Staphylococcus aureus ATCC 25923, ATCC 29213 and 30 clinical isolates of Staph. aureus were inhibited by 2-hydroxy- N -(3,4-dimethyl-5-isoxazolyl)-1,4-naphthoquinone-4-imine (I). This compound diminished bloodstream infection of mice injected i.m. with Staph. aureus; septicaemia decayed significantly when I was applied at the beginning of the infection while when I was given 3 d after bacterial challenge, a significant protection was afforded. Bactericidal activity in serum increased during the 5 h after I was administered i.p.
The acetyl derivative of I had a high MIC but when inoculated orally in mice decreased the Staph. aureus counts in circulation. This protection occurred only when the schedule of administration started close to the bacterial challenge. Antibiotic activity in vivo may be associated with in vitro effects.     

Slow heat rate increases yeast thermotolerance by maintaining plasma membrane integrity.

Thermal resistance of Saccharomyces cerevisiae was found to be drastically dependent on the kinetics of heat perturbation. Yeasts were found to be more resistant to a plateau of 1 h at 50 degrees C after a slope of temperature increase (slow and linear temperature increments) than after a shock (sudden temperature change). Thermotolerance was mainly acquired between 40-50 degrees C during a heat slope, i.e., above the maximal temperature of growth. The death of the yeasts subjected to a heat shock might be related to the loss of membrane integrity: intracellular contents extrusion, i.e., membrane permeabilization, was found to precede cell death. However, the permeabilization did not precede cell death during a heat slope and, therefore, membrane permeabilization was a consequence rather than a cause of cell death. During a slow temperature increase, yeasts which remain viable may have time to adapt their plasma membrane and thus maintain membrane integrity.     

Receptor-binding and down-regulatory properties of 22000-Mr human growth hormone and its natural 20000-Mr variant on IM-9 human lymphocytes.

Our earlier binding studies of the 22000- and 20000-Mr variants of human growth hormone (somatotropin) to pregnant-rabbit liver and mammary receptors [Closset, Smal, Gomez & Hennen (1983) Biochem. J. 214, 885-892] suggested that the 20000-Mr variant was a lower-affinity analogue of the 22000-Mr molecule. Since the receptor population in these tissues is not fully characterized, we have now investigated the binding of both variants to the well-characterized and highly specific human-growth-hormone receptor of the human lymphocyte IM-9 cell line. The maximum bindability of radioiodinated 22000- and 22000-Mr to IM-9 cells was 60 and 45% respectively. Both hormone variants have essentially the same binding characteristics: slow association (equilibrium reached in 8-10h at 30 degrees C), poor reversibility ('tight binding'), linear Scatchard plot, same specificity as shown by lack of competition by bovine, porcine or equine growth hormones or human growth hormone-(32-46)-(missing in the 20000-Mr variant),-(1-134)- and -(141-191)-peptides. Both unlabelled hormones inhibit binding of both tracers completely, with the 20000-Mr variant being only half as potent as the 22000-Mr one. The apparent affinity is 2.8 X 10(9)M-1 for the 22000-Mr variant and 1.6 X 10(9)M-1 for the 20000-Mr variant. This decreased affinity of the 20000-Mr variant appears to be due to a lower association rate constant. Concentrations (5 ng/ml) of the two variants that occupy about 15% of the total sites induce a marked down-regulation of the receptors after 18h incubation, but the 20000-Mr variant (50% decrease) has a smaller effect than the 22000-Mr variant (75% decrease). Thus the only consequence of the residues-32-46 deletion in the 20000-Mr variant is a lower association rate and affinity for the IM-9 lymphocyte human-growth-hormone receptor. The close binding characteristics of the two forms suggest that the known differences in their insulin-like effects cannot be explained by differences in the nature of their interaction with the human-growth-hormone receptor.     

A new species of bound ubisemiquinone anion in QH2: cytochrome c oxidoreductase

Using a combination of EPR and low temperature diffuse reflectance spectroscopy, a new species of semiquinone anion has been detected in QH2:cytochrome c oxidoreductase in submitochondrial particles under conditions of oxidant-induced extra reduction of cytochrome b. In contrast to the previously detected semiquinone anion, this new species is insensitive to antimycin but sensitive to treatment with 2,3-dimercaptopropanol and O2. The two species can easily be distinguished on the basis of their respective EPR properties since they differ in g-value, line width, and microwave power saturation behavior. It is concluded that the two species of semiquinone anion are bound to different domains on QH2:cytochrome c oxidoreductase. The existence of two different semiquinone anions in the enzyme strongly supports a mechanism of electron flow as proposed in the Q-cycle.     

Immunolocalization of β-1-4-galactan and its relationship with lignin distribution in developing compression wood of Cryptomeria japonica

Compression wood (CW) contains higher quantities of β-1-4-galactan than does normal wood (NW). However, the physiological roles and ultrastructural distribution of β-1-4-galactan during CW formation are still not well understood. The present work investigated deposition of β-1-4-galactan in differentiating tracheids of Cryptomeria japonica during CW formation using an immunological probe (LM5) combined with immunomicroscopy. Our immunolabeling studies clearly showed that differences in the distribution of β-1-4-galactan between NW (and opposite wood, OW) and CW are initiated during the formation of the S₁ layer. At this stage, CW was strongly labeled in the S₁ layer, whereas no label was observed in the S₁ layer of NW and OW. Immunogold labeling showed that β-1-4-galactan in the S₁ layer of CW tracheids significantly decreased during the formation of the S₂ layer. Most β-1-4-galactan labeling was present in the outer S₂ region in mature CW tracheids, and was absent in the inner S₂ layer that contained helical cavities in the cell wall. In addition, delignified CW tracheids showed significantly more labeling of β-1-4-galactan in the secondary cell wall, suggesting that lignin is likely to mask β-1-4-galactan epitopes. The study clearly showed that β-1-4-galactan in CW was mainly deposited in the outer portion of the secondary cell wall, indicating that its distribution may be spatially consistent with lignin distribution in CW tracheids of Cryptomeria japonica.