Computer simulation of polypeptides in a confinement

A coarse-grained model of polypeptide chains confined in a slit formed by two parallel impenetrable surfaces was studied. The chains were flexible heteropolymers (polypeptides) built of two kinds of united atoms—hydrophobic and hydrophilic. The positions of the united atoms were restricted to the vertices of a [310] lattice. The force field consisted of a rigorous excluded volume, a long-distance potential between a pair of amino-acid residues and a local preference for forming secondary structure (helices). The properties of the chains were studied at a wide range of temperatures from good to bad solvent conditions. Monte-Carlo simulations were carried out using the algorithm based on the chain’s local changes of conformation and employing the Replica Exchange technique. The influence of the chain length, the distances between the confining surfaces, the temperature and the force field on the dimension and the structure of chains were studied. It was shown that the presence of the confinement chain complicates the process of the chain collapse to low-temperature structures. For some conditions, one can find a rapid decrease of chain size and a second transition indicated by the rapid decrease of the total energy of the system. Figure A scheme of a polypeptide chain built on a [310] lattice and confined to a slit formed by a pair of parallel impenetrable surfaces Proceedings of “Modeling Interactions in Biomolecules II,” Prague, September 5th–9th, 2005.     

Frequently asked questions about in vivo chlorophyll fluorescence: practical issues

The aim of this educational review is to provide practical information on the hardware, methodology, and the hands on application of chlorophyll (Chl) a fluorescence technology. We present the paper in a question and answer format like frequently asked questions. Although nearly all information on the application of Chl a fluorescence can be found in the literature, it is not always easily accessible. This paper is primarily aimed at scientists who have some experience with the application of Chl a fluorescence but are still in the process of discovering what it all means and how it can be used. Topics discussed are (among other things) the kind of information that can be obtained using different fluorescence techniques, the interpretation of Chl a fluorescence signals, specific applications of these techniques, and practical advice on different subjects, such as on the length of dark adaptation before measurement of the Chl a fluorescence transient. The paper also provides the physiological background for some of the applied procedures. It also serves as a source of reference for experienced scientists.     

Synergids and filiform apparatus in the sexual and apomictic dandelions from section Palustria (Taraxacum, Asteraceae)

An evolutionary trend to reduce “unnecessary costs” associated with the sexual reproduction of their amphimictic ancestors, which may result in greater reproductive success, has been observed among the obligatory apomicts. However, in the case of the female gametophyte, knowledge about this trend in apomicts is not sufficient because most of the ultrastructural studies of the female gametophyte have dealt with amphimictic angiosperms. In this paper, we tested the hypothesis that, in contrast to amphimictic plants, synergids in apomictic embryo sacs do not form a filiform apparatus. We compared the synergid structure in two dandelions from sect. Palustria: the amphimictic diploid Taraxacum tenuifolium and the apomictic tetraploid, male-sterile Taraxacum brandenburgicum. Synergids in both species possessed a filiform apparatus. In T. brandenburgicum, both synergids persisted for a long time without any degeneration, in spite of the presence of an embryo and endosperm. We propose that the persistent synergids in apomicts may play a role in the transport of nutrients to the embryo.     

The evaluation of human papillomavirus and p53 gene mutation in benign and malignant conjunctiva and eyelid lesions

Papillomas and squamous cell carcinomas are the most common conjunctival and eyelid lesions. The etiology is still unclear and recently human papillomavirus infection and p53 gene mutation have been taken into consideration. The aim of our study was the evaluation of HPV DNApresence and p53 gene mutation in 45 benign and 38 malignant squamous lesions of the conjunctiva and eyelid. For HPV detection PCR-RFLP and immunohistochemical reaction were used; for p53 gene mutation PCR-SSCP was used. Only 8.8% papillomas, 9.1% squamous cell cancers and 3.7% basal cell cancers (using PCR-RFLP method) and 26.6% papillomas, 7.4% squamous cell cancers and 9.1% basal cell cancers (using immunohisto-chemical reaction) were HPV positive. p53 gene mutation was evaluated in 24.4% papillomas, 54.5% squamous cell cancers and 22.2% basal cell cancers; most commonly in 6 and 7 exon. Human papillomavirus infection, opposite to p53 gene mutation, is not a significant etiological factor of the benign and malignant conjunctival and eyelid lesions development.     

Flavonoids from Pinus sylvestris needles and their variation in trees of different origin grown for nearly a century at the same area

Flavonoids in needles of Scots pine planted in 1912–1914 in Poland from seeds originating from different parts of Europe, were isolated, chemically characterised and analysed by HPLC. It was shown that flavonoid profiles were similar in all tested populations and were different from those previously reported for Scots pine seedlings. They included taxifolin, taxifolin 3′-O-glucoside, quercetin as well as quercetin 3-O-glucoside and 3′-O-glucoside. The quercetin 3-O-glucoside could be found only in a trace amount in all samples and quercetin 3′-O-glucoside appeared in all samples regardless their origin. The relative concentration of taxifolin 3′-O-glucoside, quercetin, taxifolin and total flavonoids showed dependence on the origin of seeds; needles from high latitude populations contained smaller amounts of these compounds. Presented data clearly indicate that Scots pine contain glycosidases specific for glycosylation at C-3′ rather than at C-3. Besides, they indicate that long lasting influence of similar environmental factors is not able to change genetic regulatory systems responsible for flavonoid biosynthesis.     

Timing of the first zygotic cleavage as a marker of developmental potential of mammalian embryos

Embryo quality related to its developmental potential is now one of the most important issues in modern embryology. It has been demonstrated that some in vitro produced blastocysts fail to hatch and implant after transfer despite a normal morphology. Although embryos are able to adjust to sub-optimal culture conditions, significant changes in expression profiles of developmentally important genes have been noticed. Timing of the first zygotic cleavage is considered a non-invasive marker of embryo developmental potential and has been successfully used in human IVF programs for identifying embryos of superior quality. Early-cleaving zygotes are more likely to develop to the blastocyst stage than their late-cleaving counterparts. The timing of the first zygotic cleavage has been associated with several parameters that may affect developmental potential of the resulting embryos. The mechanism causing variation in the timing of the first zygotic cleavage has not been identified. It may be related to culture environment or to some intrinsic factors within the oocyte, the sperm or both. In this paper we discuss some of the important aspects related to the timing of the first zygotic cleavage and its influence on the developmental competence of resulting embryos.     

Usefulness of copeptin as a potential biomarker in TBE

Objective: The aim of the study was to evaluate the usefulness of copeptin for differentiation of hyponatremia in the course of tick-borne encephalitis (TBE) and for being a prognostic marker of the severity of TBE.

Materials and methods: One hundred and fourteen patients with TBE were included in the study. The control group consisted of 62 patients diagnosed with viral meningitis.

Results: Copeptin concentration did not differ in patients with hyponatremia and normonatremia. Urinary sodium excretion to plasma copeptin (copeptin/UNa Secretion) ratio was significantly lower in Syndrome of Inappropriate Antidiuretic Hormone (SIADH) Secretion patients than in patients with hyponatremia of other origin. Mean copeptin concentration in TBE patients was higher than in control group (VM) patients. There were no differences between patients with severe and mild course of TBE.

Conclusions: Copeptin/UNa ratio may be used as a potential biomarker of SIADH in patients with TBE. Copeptin concentration is significantly higher in patients with TBE than in viral meningitis of other origin, especially in patients aged 18–34 and >49 years old. Copeptin does not differentiate TBE of mild and severe course.     

Putative Structural and Functional Coupling of the Mitochondrial BKCa Channel to the Respiratory Chain

Potassium channels have been found in the inner mitochondrial membranes of various cells. These channels regulate the mitochondrial membrane potential, the matrix volume and respiration. The activation of these channels is cytoprotective. In our study, the single-channel activity of a large-conductance Ca²⁺-regulated potassium channel (mitoBK_Ca channel) was measured by patch-clamping mitoplasts isolated from the human astrocytoma (glioblastoma) U-87 MG cell line. A potassium-selective current was recorded with a mean conductance of 290 pS in symmetrical 150 mM KCl solution. The channel was activated by Ca²⁺ at micromolar concentrations and by the potassium channel opener NS1619. The channel was inhibited by paxilline and iberiotoxin, known inhibitors of BK_Ca channels. Western blot analysis, immuno-gold electron microscopy, high-resolution immunofluorescence assays and polymerase chain reaction demonstrated the presence of the BK_Ca channel β4 subunit in the inner mitochondrial membrane of the human astrocytoma cells. We showed that substrates of the respiratory chain, such as NADH, succinate, and glutamate/malate, decrease the activity of the channel at positive voltages. This effect was abolished by rotenone, antimycin and cyanide, inhibitors of the respiratory chain. The putative interaction of the β4 subunit of mitoBK_Ca with cytochrome c oxidase was demonstrated using blue native electrophoresis. Our findings indicate possible structural and functional coupling of the mitoBK_Ca channel with the mitochondrial respiratory chain in human astrocytoma U-87 MG cells.     

Structural integrity of the PCI domain of eIF3a/TIF32 is required for mRNA recruitment to the 43S pre-initiation complexes

Transfer of genetic information from genes into proteins is mediated by messenger RNA (mRNA) that must be first recruited to ribosomal pre-initiation complexes (PICs) by a mechanism that is still poorly understood. Recent studies showed that besides eIF4F and poly(A)-binding protein, eIF3 also plays a critical role in this process, yet the molecular mechanism of its action is unknown. We showed previously that the PCI domain of the eIF3c/NIP1 subunit of yeast eIF3 is involved in RNA binding. To assess the role of the second PCI domain of eIF3 present in eIF3a/TIF32, we performed its mutational analysis and identified a 10-Ala-substitution (Box37) that severely reduces amounts of model mRNA in the 43–48S PICs in vivo as the major, if not the only, detectable defect. Crystal structure analysis of the a/TIF32-PCI domain at 2.65-Å resolution showed that it is required for integrity of the eIF3 core and, similarly to the c/NIP1-PCI, is capable of RNA binding. The putative RNA-binding surface defined by positively charged areas contains two Box37 residues, R363 and K364. Their substitutions with alanines severely impair the mRNA recruitment step in vivo suggesting that a/TIF32-PCI represents one of the key domains ensuring stable and efficient mRNA delivery to the PICs.