His-tags as Zn(II) binding motifs in a protein-based fluorescent sensor

Fluorescent indicators that allow real-time imaging of Zn(II) in living cells are invaluable tools for understanding Zn(II) homeostasis. Genetically encoded sensors based on fluorescence resonance energy transfer between fluorescent protein domains have important advantages over synthetic probes. We discovered that hexahistidine tags have a strong tendency to dimerize upon binding of Zn(II) in solution and we used this principle to develop a new protein-based sensor for Zn(II). Enhanced cyan and yellow fluorescent proteins were connected by long flexible peptide linkers and His-tags were incorporated at both termini of this fusion protein. The resulting sensor CLY9-2His allows the ratiometric fluorescent detection of Zn(II) in the nanomolar range. In addition, CLY9-2His is selective over the physiologically relevant metal ions Fe(II), Mn(II), Ca(II) and Mg(II). Our approach demonstrates the potential of using small peptides as metal-binding ligands in chelating fluorescent protein chimeras.     

Feeding patterns and weight among First Nations children.

PURPOSE: Little information is available on the growth and feeding patterns of First Nations children. Our goal was to assess the anthropometric status, feeding practices, and dietary intake of children born in 1994 or 1995 (n=102) and living in Walpole Island First Nation. METHODS: Information on demographic characteristics and infant feeding practices was obtained through parent interviews conducted between 1994 and 1999. Parents also completed a 24-hour dietary recall for their children when they were 48 months old. Head circumference was measured at three and 18 months, and weight and length/height at three, 18, 33, and 48 months. RESULTS: Most infants (75%) were breastfed at birth; however, by the time infants were three months old, 39.7% of the mothers had stopped breastfeeding. Over half (57.1%) of the infants were fed solids before age three months, 11.6% were given whole milk before age nine months, and 59.4% had low fat milk before age two years. Body mass index (kg/m2) (BMI) was above the 85th percentile for 27.8% at the 48-month interview, and was associated with a maternal BMI above 25 (OR=7.8, CI=1.1-41.9). CONCLUSIONS: Mothers need to be encouraged to follow current infant feeding recommendations. Strategies should be developed to reduce the prevalence of overweight among adults and children in First Nations communities.     

Direct determination of fat using ISO/CD 17189.2—an international collaborative study of spreadable fats and a meta-analysis

A method for the direct determination of the total fat content in spreadable fats, including low fat and regular fat margarines, reduced fat butter, blended spreads and a sterol-fortified spread, was studied collaboratively using 12 laboratories and eight samples (fat range: 23–82 g fat/100 g product). The method involved mixing 1–2 g of sample with petroleum ether solvent to extract the fat. The solvent of pooled lots of 3–4 extractions (the number of extractions depended on the phase separation technique used) was evaporated, and the extract was dried to constant mass and weighed. Estimates of repeatability (2.8s_r) and reproducibility (2.8s_R) were 0.24 g fat/100 g product and 0.44 g fat/100 g product, respectively. A meta-analysis of five international collaborative studies yielded grand estimates of repeatability and reproducibility of 0.26 g fat/100 g product and 0.45 g fat/100 g product, respectively (95% confidence intervals: 0.24–0.28 g fat/100 g product and 0.41–0.49 g fat/100 g product, respectively). These estimates apply to the full scope of draft ISO standard 17189.2, i.e. butter, edible oil emulsions and spreadable fats. The work reported in this paper completes the validation of this standard, which may now be progressed for final adoption as an international reference method.     

Scanning Electron Microscopy of Wheat Starch. IV. Digestion of large Granules by Glucoamylase of Fungal (Aspergillus niger) Origin)

Starch granules attacked by glucoamylase of fungal origin show a pattern of erosion which is different from that resulting from wheat alpha-amylase attack. The glucoamylase pattern, which is characterised by a relatively uniform erosion of the surface is discussed in comparison with that of alpha-amylase and possible reasons for the differences are suggested.     

Scanning Electron Microscopy of Wheat Starch. III. Granule Development in the Endosperm

Study of A type starch granules from immature wheat endosperm with the scanning electron microscope has led to a developmental sequence being suggested. Evidence is provided which indicates that the initial minute spherical granule becomes a “nucleus” which is progressively surrounded in the equatorial plane by a continually enlarging structure which resembles two lips with a furrow between them. When the lips have completely surrounded the “nucleus” the thickness and diameter of the granule continue to increase in such a way that the furrow becomes shallower and is present in the mature biconvex structure only as a shallow score.     

Reaktionen mit Thio-bis-formamidinen

Reactions with Thio-bis-formamidines N,N,N′-Trisubstituted S-(4-amino-s-triazin-2-yl)-isothioureas 3 are prepared in good yields form trisubstituted thio-bis-formamidine-hydrochlorides 1 and esters of cyanimidocarboxylic acids 2 . In aqueous alkali solutions compounds 1 react with β-dicarbonyl compounds under rearrangement to yield substituted N-(pyrimidin-2-yl)-thioureas 4 and 5 .     

Genetics of migraine

Several historical reports focusing on the heredity of migraine, as well as recent studies on its epidemiology and molecular biology, have revealed evidence for a decisive role of genetic factors in the aetiopathogenesis of familial migraine. Indeed, family studies, segregation analyses and twin studies have shown that genetic factors play an important role in disposition towards migraine but could not explain the entire aetiopathogenesis. The influence of extragenetic factors, however, remains mostly unknown. Recent linkage analyses have provided evidence for genetic heterogeneity. A locus for Familial Hemiplegic Migraine (FHM), the only known type of migraine that follows autosomaldominant transmission, has been linked to chromosome 19p13 but genetic heterogeneity has also been shown, i.e., different types of migraine could be excluded from this locus. Further investigations should concentrate on identifying the FHM gene on chromosome 19p13, on linkage analyses with markers for different susceptibility genes, and on genomic analyses of highly informative pedigrees. This would lead to further clues to the pathogenesis underlying migraine and, thus, to therapeutic developments.     

Conditions, interventions, and outcomes in nursing research: a comparative analysis of North American and European/International journals. (1981-1990)

This study compared the conceptual foci and methodological characteristics of research projects which tested the effects of nursing interventions, published in four general nursing research journals with predominantly North American, and two with predominantly European/International authorship and readership. Dimensions and variables of comparison included: nature of subjects, design issues, statistical methodology, statistical power, and types of interventions and outcomes. Although some differences emerged, the most striking and consistent finding was that there were no statistically significant differences (and thus similarities) in the content foci and methodological parameters of the intervention studies published in both groups of journals. We conclude that European/International and North American nursing intervention studies, as reported in major general nursing research journals, are highly similar in the parameters studied, yet in need of overall improvement. Certainly, there is no empirical support for the common (explicit or implicit) ethnocentric American bias that leadership in nursing intervention research resides with and in the United States of America.     

Assembly of alcohol oxidase in peroxisomes of the yeast Hansenula polymorpha requires the cofactor flavin adenine dinucleotide

The peroxisomal flavoprotein alcohol oxidase (AO) is an octamer (600 kDa) consisting of eight identical subunits, each of which contains one flavin adenine dinucleotide molecule as a cofactor. Studies on a riboflavin (Rf) auxotrophic mutant of the yeast Hansenula polymorpha revealed that limitation of the cofactor led to drastic effects on AO import and assembly as well as peroxisome proliferation. Compared to wild-type control cells Rf-limitation led to 1) reduced levels of AO protein, 2) reduced levels of correctly assembled and activated AO inside peroxisomes, 3) a partial inhibition of peroxisomal protein import, leading to the accumulation of precursors of matrix proteins in the cytosol, and 4) a significant increase in peroxisome number. We argue that the inhibition of import may result from the saturation of a peroxisomal molecular chaperone under conditions that normal assembly of a major matrix protein inside the target organelle is prevented.