Compensating bends in a 16-base-pair DNA oligomer containing a T(3)A(3) segment: A NMR study of global DNA curvature

In-phase ligated DNA containing T(n)A(n) segments fail to exhibit the retarded polyacrylamide gel electrophoresis (PAGE) migration observed for in-phase ligated A(n)T(n) segments, a behavior thought to be correlated with macroscopic DNA curvature. The lack of macroscopic curvature in ligated T(n)A(n) segments is thought to be due to cancellation of bending in regions flanking the TpA steps. To address this issue, solution-state NMR, including residual dipolar coupling (RDC) restraints, was used to determine a high-resolution structure of [d(CGAGGTTTAAACCTCG)2], a DNA oligomer containing a T3A3 tract. The overall magnitude and direction of bending, including the regions flanking the central TpA step, was measured using a radius of curvature, Rc, analysis. The Rc for the overall molecule indicated a small magnitude of global bending (Rc = 138 +/- 23 nm) towards the major groove, whereas the Rc for the two halves (72 +/- 33 nm and 69 +/- 14 nm) indicated greater localized bending into the minor groove. The direction of bending in the regions flanking the TpA step is in partial opposition (109 degrees), contributing to cancellation of bending. The cancellation of bending did not correlate with a pattern of roll values at the TpA step, or at the 5' and 3' junctions, of the T3A3 segment, suggesting a simple junction/roll model is insufficient to predict cancellation of DNA bending in all T(n)A(n) junction sequence contexts. Importantly, Rc analysis of structures refined without RDC restraints lacked the precision and accuracy needed to reliably measure bending.     

Midgut glycosidases activities in monophagous larvae of Apollo butterfly, Parnassius apollo ssp. frankenbergeri

Parnassius apollo (Lepidoptera, Papilionidae) declines on numerous localities all over Europe. Its local subspecies frankenbergeri, inhabiting the Pieniny Mts (southern Poland) and successfully recovered from extinction, is monophagous in larval stage. In natural conditions, it completes development on the orpine Sedum telephium ssp. maximum. Since proper quality and quantity of necessary nutritional compounds of the food plant ensure developmental success, the digestive processes in the insect midgut should reflect adaptation to a specific food source. The paper presents, for the first time, the activity of detected glycolytic enzymes in midgut tissue and liquid gut contents of the L4 and L5 instars of P. apollo larvae. alpha-Amylase plays the main role in utilization of carbohydrates, contrary to cellulase activity. Saccharase seems to be the main disaccharidase, and high activity of beta-glycosidase enables hydrolysis of the plant glycosides. Trehalase activity was unexpectedly low and comparable to those of cellobiase and lactase. alpha-Amylolytic and other glycolytic activities indicate that larvae utilize starch and other carbohydrate compounds as energy sources. Possible use of some plant allelochemicals as energy sources by Apollo larvae is discussed.     

Genetic variation in the efficiency of nitrogen utilization and photosynthetic activity of flag leaves among the old and modern germplasm of winter wheat

Genotypic variation in major components of the efficiency of nitrogen utilization and photosynthetic activity of flag leaves among old (released 1881-1963) and modern (released 1969-2003) cultivars of winter wheat was studied in field conditions under varied N fertilization levels (110, 90 and 80 kg N ha-1). Significant genotypic differences were observed for all characters. Their heritabilities ranged from 0.37 to 0.93 and were the lowest for the leaf efficiency of gas exchange, photosynthetic rate, straw N content and the economic index of N utilization efficiency (NUE). Some modern cultivars exhibited an enhanced tolerance to N shortage and several attributes of efficient N utilization (e.g. later senescing and more photosynthetically active flag leaves, increased ability to redistribute N into grains). The genotypes may serve as donors of appropriate characteristics for breeding. The observed cultivar-by-fertilization interactions suggest, however, that evaluations under diverse fertilization regimes may be necessary when searching for improved wheat efficiency and adaptation to less favourable environments.     

Oxidatively modified proteins and DNA in digestive gland cells of the fresh-water mussel Unio tumidus in the presence of tannic acid and its derivatives

The oxidative effect of tannic acid and its two derivatives (ellagic and gallic acid), naturally occurring plant polyphenols, has been studied on digestive gland cells of the fresh-water mussel Unio tumidus. A spectrophotometric method was used to determine the protein thiol groups after incubation of the cells with the polyphenols at concentrations of 1, 15 and 60 microM. The results showed that the oxidative modification of proteins increased in a concentration-dependent manner but no changes were observed at the concentration of 1 microM. The comet assay (single-cell gel electrophoresis assay) with the formamido-pyrimidine glycosylase (FPG) protein was used to assess oxidative DNA base damage. The cells were treated with polyphenols at the concentrations of 30 and 60 microM and post-incubated with FPG. FPG strongly enhanced DNA damage induced by the polyphenols, indicating that N-7 guanine oxidation is responsible for the observed effect. Using the comet assay in combination with proteinase K we were able to demonstrate the presence of DNA-protein cross-links as the probable cause of the decrease in DNA migration. After treatment of the cells with tannic acid and its metabolites at concentrations of 120, 180 and 240 microM, they were post-incubated with proteinase K. After this treatment an increased DNA migration was observed, indicating the presence of DNA-protein cross-links. We have also used a fluorescence method with Hoechst 33258/propidium iodide DNA-binding dyes to study the extent of DNA fragmentation after exposure of the cells to polyphenols at concentrations of 1, 5 and 60 microM. The results demonstrate that the polyphenols can induce apoptosis and necrosis at higher concentrations (5 and 60 microM). All experimental data suggest that tannic, ellagic and gallic acids at concentrations above 1 microM are able to interact with proteins and DNA, which leads to their degradation or changes in their function.