The influence of the rolC gene on isoflavonoid production in callus cultures of Maackia amurensis

The polyphenolic complex of Maackia amurensis, as well as a complex of isoflavonoids from M. amurensis callus cultures, display strong hepatoprotective effects in experimental animal and human studies. To increase the yield of polyphenols in cultures of M. amurensis, calli were transformed with the rolC gene as well as with an empty vector that was used as a control. HPLC analysis revealed that the transgenic cultures produced the same complex of isoflavonoids. The complex consisted of 20 compounds, including isoflavones and their glucosides as well as pterocarpans and their glucosides. The cultures transformed with either the empty vector or the rolC gene construct produced on average 1.22 % dry weight (DW) and 1.39 % DW of isoflavonoids, respectively. Isoflavonoid production in the transformed callus lines carrying the empty vector and the rolC gene construct reached 106 and 146 mg/L, respectively. Moreover, the rolC gene construct promoted cell growth and overall cell productivity. The transgenic callus lines expressing the rolC gene exhibited higher levels of the following six isoflavonoids: daidzein, calycosin, formononetin, 4′-Ο-β-glucopyranosyldaidzin, maackiain and 6′-O-malonyl-3-O-β-D-glucopyranosylmaackiain. However, lower levels of genistin were observed in rolC calli than in those carrying the empty vector.     

Spatial structure and antigenic activity of non-glycosylated forms of embryonal prealbumin-1 (EPA-1NG)]

The influence of temperature and pH on the spatial structure of EPA-1NG has been studied by means of circular dichroism and differential UV-spectroscopy, indicating the molecule to consist mainly of beta-structures. A conformational transition in the molecule was observed within the range of 40-50 degrees C. The further temperature elevation (up to 70 degrees C) was accompanied to the complete distortion of the parent conformation, which is reversed after cooling down to 20 degrees C. A correlation of the spectral data with the antigenic activity of genuine EPA-1NG and its carboxymethylated, heat-degraded and pH-denatured derivatives demonstrates that some antigenic determinants of EPA-1NG appear to be topographic.     

Cyclic nucleotide specificity of the activator and catalytic sites of a cGMP-stimulated cGMP phosphodiesterase from Dictyostelium discoideum

The cellular slime mold Dictyostelium discoideum has an intracellular phosphodiesterase which specifically hydrolyzes cGMP. The enzyme is activated by low cGMP concentrations, and is involved in the reduction of chemoattractant-mediated elevations of cGMP levels. The interaction of 20 cGMP derivatives with the activator site and with the catalytic site of the enzyme has been investigated. Binding of cGMP to the activator site is strongly reduced (more than 80-fold) if cGMP is no longer able to form a hydrogen bond at N2H2 or O2'H. Modifications at N7, C8, O3' and O5' induce only a small reduction of binding affinity. A cyclic phosphate structure, as well as a negatively charged oxygen atom at phosphorus, are essential to obtain activation of the enzyme. Substitution of the axial exocyclic oxygen atom by sulphur is tolerated; modification of the equatorial oxygen atom reduces the binding activity of cGMP to the activator site by 90-fold. Binding of cGMP to the catalytic site is strongly reduced if cGMP is modified at N1H, C6O, C8 and O3', while modifications at N2H2, N3, N7, O2'H, and O5' have minor effects. Both exocyclic oxygen atoms are important to obtain binding of cGMP to the catalytic site. The results indicate that activation of the enzyme by cGMP and hydrolysis of cGMP occur at different sites of the enzyme. cGMP is recognized at these sites by different types of molecular interaction between cGMP and the protein. cGMP derivatives at concentrations which saturate the activator site do not induce the same degree of activation of the enzyme (activation 2.3-6.6-fold). The binding affinities of the analogues for the activator site and their maximal activation are not correlated. Our results suggest that the enzyme is activated because cGMP bound to the activator site stabilizes a state of the enzyme which has a higher affinity for cGMP at the catalytic site.     

Critical analysis of protein signaling networks involved in the regulation of plant secondary metabolism: focus on anthocyanins

Anthocyanin biosynthesis in Arabidopsis is a convenient and relatively simple model for investigating the basic principles of secondary metabolism regulation. In recent years, many publications have described links between anthocyanin biosynthesis and general defense reactions in plants as well as photomorphogenesis and hormonal signaling. These relationships are complex, and they cannot be understood intuitively. Upon observing the lacuna in the Arabidopsis interactome (an interaction map of the factors involved in the regulation of Arabidopsis secondary metabolism is not available), we attempted to connect various cellular processes that affect anthocyanin biosynthesis. In this review, we revealed the main signaling protein modules that regulate anthocyanin biosynthesis. To our knowledge, this is the first reconstruction of a network of proteins involved in plant secondary metabolism.     

Possibility of reversing the action of the membranotropic antibiotic, gramicidin S, on bacteria

The study on the possibility of eliminating gramicidin S from the bacterial cells which had adsorbed it showed that a part of the labeled antibiotic bound by the bacteria may be washed out with buffer or salines. When the cells which had adsorbed gramicidin S were treated with lecithin emulsion, a significant part of the bound antibiotic was transferred to the lecithin liposomes. This turned the gramicidin S effect to the cells: significant but not complete reduction of the membrane barrier properties and dehydrogenase reactivation. Elimination of gramicidin S also reduced the colony forming capacity in a part of the cells.     

Slow plasma waves in a fine-stratified periodic semiconductor structure

A study is made of the propagation of electromagnetic waves in an infinite fine-stratified periodic structure composed of alternating layers of two different semiconductors. Such a periodic structure, which can be treated as a uniform uniaxial semiconductor with two different values of the dielectric constant, has four characteristic frequencies: two plasma frequencies of the layers and two hybrid frequencies, which are determined by the parameters of the layers. These frequencies govern the dispersion properties of the eigenwaves of the semiconductor structure. The effect of the dissipative processes on the wave dispersion is considered, and the dependence of the minimum phase velocity on the thickness of the layers and on the collision frequency between charge carriers within them is determined.     

The state of lipid peroxidation and enzymes of calcium transport system in sarcoplasmic reticulum of ischemic myocardium

In 30 experiments on mongrel dog hearts it was shown that 30 min of total ischemia (37 degrees C) followed by accumulation of MDA in the SR membranes and decrease of their Ca2+-uptake, but had no effect on activity Ca2+-ATPase. After 60-120 min ischemia marked a decrease of Ca2+-uptake and activity Ca2+-ATPase took place, MDA content remained at the increased level. The results show that lipid peroxidation take part in the increase of the permeability of SR membranes for Ca2+ and inhibiting of Ca2+-ATPase.