The DCX-domain tandems of doublecortin and doublecortin-like kinase

The doublecortin-like domains (DCX), which typically occur in tandem, are novel microtubule-binding modules. DCX tandems are found in doublecortin, a 360-residue protein expressed in migrating neurons; the doublecortin-like kinase (DCLK); the product of the RP1 gene that is responsible for a form of inherited blindness; and several other proteins. Mutations in the gene encoding doublecortin cause lissencephaly in males and the 'double-cortex syndrome' in females. We here report a solution structure of the N-terminal DCX domain of human doublecortin and a 1.5 A resolution crystal structure of the equivalent domain from human DCLK. Both show a stable, ubiquitin-like tertiary fold with distinct structural similarities to GTPase-binding domains. We also show that the C-terminal DCX domains of both proteins are only partially folded. In functional assays, the N-terminal DCX domain of doublecortin binds only to assembled microtubules, whereas the C-terminal domain binds to both microtubules and unpolymerized tubulin.     

Acid-base equilibria at interface separating electrolyte solution and lipid bilayer formed from phosphatidylcholine

The dependence of the interfacial tension of a lipid membrane formed from phosphatidylcholine on the pH of the aqueous solution has been studied. The model describing the H(+) and OH(-) ions adsorption in the bilayer lipid surface has been presented in this work. We take suitable equations to describe the dependence of interfacial tension of a lipid bilayer membrane on H(+) and OH(-) ion concentrations. A theoretical equation is derived to describe this dependence in the whole pH range.     

Conformational Studies of PACAP(1–27) and Its Fragments

Summary Conformational features of the neuropeptide pituitary adenylate cyclase activating polypeptide (1–27) (PACAP(1–27)) and its shorter fragments (1–5), (7–11) and (14–27) were studied by circular dichroism (CD) and fluorescence spectroscopy. The obtained CD spectra revealed that only PACAP(1–27) and the fragment (14–27) possess some content of an organized structure — the α-helix. This C-terminal, helical part of the peptides is important for receptor binding as it provides a stable structure that can reside in the ordered lipid region of the receptor site in the membrane, while the primary biological function of the hormone resides in the N-terminal, disordered part. Fluorescence studies have revealed that the tyrosine residue located in the helical region of PACAP has a higher quantum yield and a longer average lifetime than the tyrosine in the N-terminus, probably due to a ‘shielding’ effect of the hydrophobic cluster around Tyr²².     

Differences in Energy Balance-Related Behaviours in European Preschool Children: The ToyBox-Study

Background

The aim of the current study was to compare levels of energy balance-related behaviours (physical activity, sedentary behaviour, and dietary behaviours (more specifically water consumption, sugar-sweetened beverage consumption and unhealthy snacking)) in four- to six-year-old preschoolers from six European countries (Belgium, Bulgaria, Germany, Greece, Poland, and Spain) within the ToyBox cross-sectional study.

Methods

A sample of 4,045 preschoolers (4.77 ± 0.43 years; 52.2% boys) had valid physical activity data (steps per day), parents of 8,117 preschoolers (4.78 ± 0.46 years; 53.0% boys) completed a parental questionnaire with questions on sedentary behaviours (television viewing, computer use, and quiet play), and parents of 7,244 preschoolers (4.77 ± 0.44 years; 52.0% boys) completed a food frequency questionnaire with questions on water consumption, sugar-sweetened beverage consumption and unhealthy snacking.

Results

The highest levels of physical activity were found in Spain (12,669 steps/day on weekdays), while the lowest levels were found in Bulgaria and Greece (9,777 and 9,656 steps/day on weekdays, respectively). German preschoolers spent the least amount of time in television viewing (43.3 min/day on weekdays), while Greek preschoolers spent the most time in television viewing (88.5 min/day on weekdays). A considerable amount of time was spent in quiet play in all countries, with the highest levels in Poland (104.9 min/day on weekdays), and the lowest levels in Spain (60.4 min/day on weekdays). Belgian, German, and Polish preschoolers had the lowest intakes of water and the highest intakes of sugar-sweetened beverages. The intake of snacks was the highest in Belgian preschoolers (73.1 g/day) and the lowest in Greek preschoolers (53.3 g/day).

Conclusions

Across six European countries, differences in preschoolers’ energy balance-related behaviours were found. Future interventions should target European preschoolers’ energy balance-related behaviours simultaneously, but should apply country-specific adaptations.     

TLR9 2848 GA Heterozygotic Status Possibly Predisposes Fetuses and Newborns to Congenital Infection with Human Cytomegalovirus

Background

Some single nucleotide polymorphisms (SNP), located in Toll-like receptor (TLR) genes, were reported to be associated with human cytomegalovirus (HCMV) infections. The study was aimed to assess the correlation of SNPs at TLR4 and TLR9 genes with the occurrence of congenital cytomegaly, based on available samples.

Methods

Reported case-control study included both HCMV infected and non-infected fetuses and newborns. The specimens were classified to the molecular analyses, based on serological features of the recent infection and HCMV DNAemia in body fluids. TLR SNPs were studied, using multiplex nested PCR-RFLP assay, and determined genotypes were confirmed by sequencing. Hardy-Weinberg equilibrium was assessed for the identified genotypes. The linkage disequilibrium was also estimated for TLR4 SNPs. A relationship between the status of TLR genotypes and congenital cytomegaly development was estimated, using a logistic regression model.

Results

Hardy Weinberg equilibrium was observed for almost all SNPs, both infected and non-infected patients, with exception of TLR4 896 A>G polymorphism in the control group (P≤0.050). TLR4 896 A>G and 1196 C>T SNPs were found in linkage disequilibrium in both study groups (P≤0.050). The CC genotype at TLR4 1196 SNP and the GA variant at TLR9 2848 G>A SNP were significantly associated with HCMV infection (P≤0.050). The risk of congenital cytomegaly was higher in heterozygotes at TLR9 SNP than in the carriers of other genotypic variants at the reported locus (OR 4.81; P≤0.050). The GC haplotype at TLR4 SNPs and GCA variants at TLR4 and TLR9 SNPs were significantly associated with HCMV infection (P≤0.0001). The ACA variants were more frequent among fetuses and neonates with symptomatic, rather than asymptomatic cytomegaly (P≤0.0001).

Conclusions

TLR4 and TLR9 polymorphisms may contribute to the development of congenital infection with HCMV in fetuses and neonates. The TLR9 2848 GA heterozygotic status possibly predisposes to HCMV infection, increasing the risk of congenital cytomegaly development.     

Complex of Rutin with β-Cyclodextrin as Potential Delivery System

This study aimed to obtain and characterize an RU-β-CD complex in the context of investigating the possibility of changes in the solubility, stability, antioxidative and microbiological activity as well as permeability of complexated rutin as against its free form. The formation of the RU-β-CD complex via a co-grinding technique was confirmed by using DSC, SEM, FT-IR and Raman spectroscopy, and its geometry was assessed through molecular modeling. It was found that the stability and solubility of the so-obtained complex were greater compared to the free form; however, a slight decrease was observed inits antibacterial potency. An examination of changes in the EPR spectra of thecomplex excluded any reducing effect of complexation on the antioxidative activity of rutin. Considering the prospect of preformulation studies involving RU-β-CD complexes, of significance is also the observed possibility of prolongedly releasing rutin from the complex at a constant level over along period of 20 h, and the fact that twice as much complexated rutin was able topermeate compared to its free form.     

Methods of the site-selective solid phase synthesis of peptide-derived Amadori products

Two procedures of glycated peptides’ synthesis have been developed. The first method involves reductive alkylation of the ε-amino groups of lysine with 2,3:4,5-di-O-isopropylidene-β-d-arabino-hexos-2-ulo-2,6-pyranose in the presence of sodium cyanoborohydride on solid support. The second one uses a new fully protected lysine derivative, which is a building block designed for direct introduction of the glycated lysine moiety into a peptide, according to the standard solid phase synthesis protocol. The applicability of the proposed methods for the synthesis of peptide-derived Amadori products is discussed. The structure of the synthesized glycated peptides was confirmed by high-resolution mass spectrometry and enzymatic hydrolysis. Circular dichroism studies, performed in water solution, revealed that the formation of the Amadori rearrangement product in the lysine side chain does not influence significantly the conformational preferences of the peptides studied. However, when the solvent was changed to trifluoroethanol, the glycated peptides preferred β-turn conformation.     

Iodine, Selenium, and Other Trace Elements in Urine of Pregnant Women

The purpose of this work was to determine trace element levels in urine and evaluate possible associations between urinary iodine concentration (UIC), other trace elements (Cr, Cu, Fe, Mn, Na, Se, Zn), toxic elements (Cd, Pb), anthropometrical measures (body weight and height), glycemic indices (serum insulin and glucose), and several parameters related to thyroid function (thyroid stimulating hormone, free thyroxine, antithyroid peroxidase antibodies, thyroid volume, and thyroid echogenicity) in pregnant women. One hundred sixty-nine participants were recruited. The whole study group, originating from Krakow region, comprised three subgroups belonging to three trimesters: I trimester (n?=?28), II trimester (n?=?83), and III trimester (n?=?58). Trace elements were determined using inductively coupled plasma mass/(atomic emission) spectrometry. Partial least square model was used to reveal correlation structure between parameters investigated, as well as a possible causal relationship between dependent parameters and potentially explanatory parameters. Results obtained for trace and toxic elements in urine were comparable with results of other authors, although the study group was not homogenous. We confirmed (1) low iodine excretion in pregnant women, (2) the existence of statistically significant correlation between UIC and urinary selenium, and (3) lack of correlation between latter parameter and typical indices of thyroid function. Urinary selenium correlated with other urinary trace elements, but physiological significance of this finding remains uncertain. The fact that a large number of pregnant women fail to meet dietary recommendations for iodine is the major reason for concern.     

Polo box domain of Plk3 functions as a centrosome localization signal, overexpression of which causes mitotic arrest, cytokinesis defects, and apoptosis

Polo-like kinase 3 (Plk3), an immediate early response gene product, plays an important role in the regulation of mitosis, DNA damage checkpoint activation, and Golgi dynamics. Similar to other members of the Plk family, Plk3 has a conserved kinase domain at the N terminus and a Polo box domain consisting of two Polo boxes at the C terminus. In this study, we demonstrate that the Polo box domain of Plk3 is sufficient for subcellular localization of this kinase to the centrosomes, the spindle poles, and the midbody when ectopically expressed in HeLa and U2OS cells. Both Polo boxes are required for the subcellular localization. Overexpression of the Polo box domain, not the kinase domain, of Plk3 causes significant cell cycle arrest and cytokinesis defects, eventually leading to mitotic catastrophe/apoptosis. Interestingly, the Polo box domain of Plk3 is more potent in inhibiting cell proliferation and inducing apoptosis than that of Plk1, suggesting that this domain can provide an additional structural basis for discovery of new anticancer drugs given the current emphasis on Plk1 as a therapeutic target.