Enzyme-Complemented Activatorsorbent Assay (ECASA): Genetic Engineering for Enzyme-Linked Immunosorbent Assay-Type Mercuric Ion Detection

The sensor component of bacterial mercury resistance systems is the metalloregulatory protein MerR, which has nanomolar sensitivity and high selectivity for Hg(II). A fusion protein of MerR and the α-peptide part of β-galactosidase (LacZα) was constructed by fusing the relevant genes. The protein exhibited both MerR functions and α-complementing activity to the inactive LacZΔM15 (M15) protein. The bifunctional character of the appropriate MerR–LacZα-complemented M15 protein (MerR–LacZα:M15 protein complex) was used to develop a Hg(II)-specific enzyme-complemented activatorsorbent assay. Hg(II) was immobilized and presented on a matrix taking advantage of the high affinity of Hg(II) to SH residues. The immobilized Hg(II) could be specifically detected down to the parts-per-billion level by quantifying the β-galactosidase activity of the bound fusion protein complex.     

Synthesis and properties of sulfated alkyl glycosides

Alkyl glycosides were sulfated with sulfur trioxide-pyridine. Dodecyl - and β- -glucopyranoside gave the corresponding 6-sulfates in 75 and 51% yields, respectively. Separation from polysulfated compounds was carried out by reversed-phase HPLC. Tetradecyl β-maltopyranoside (16) gave a 88:12 mixture of 6′- and 6-sulfates. The sulfated compounds were characterized by ¹H-, ¹³C-, and 2-dimensional NMR spectroscopy. Surfactant and thermotropic liquid-crystalline properties of the sugar derivatives were examined. All of the glycosides show smectic phases (S_A), and the clearing points rise by introduction of sulfate groups. Even glycosides having no unprotected hydroxy groups may show S_A-phases when bearing sulfate groups. The mesomorphic properties cannot be explained by formation of distinct aggregates, but rather must be interpreted by an effective intramolecular contrast.     

Use of a layered double hydroxide (LDH) to buffer nitrate in soil: long-term nitrate exchange properties under cropping and fallow conditions

The potential use of a layered double hydroxide (LDH) to act as a nitrate buffer system in soil in order to reduce the movement of nitrate was investigated. Long-term plant and soil experiments were carried out under greenhouse conditions with the following objectives: (i) evaluate the nitrate adsorption capacity of the LDH during crop growth, and its influence on N uptake, (ii) study the ability of the LDH to adsorb nitrate mineralized during fallow periods, and its influence on nitrate leaching, (iii) evaluate the reversibility for nitrate exchange of the LDH under cultivation conditions, and (iv) determine the nitrate buffer capacity of the soil after LDH application. The LDH adsorbed nitrate from the soil solution during the growth period without affecting plant N uptake. As a result of the adsorption of nitrate on the LDH, the nitrate-N concentration in the soil solution at harvest was reduced by a factor of ten compared to a soil without LDH. The LDH efficiently adsorbed nitrate that was mineralized in the soil during periods without cultivation, reduced nitrate-N leaching losses by about 80%, and kept this nitrate available for a following crop. The nitrate buffer capacity of the soil after 15months increased from 0.3 (without LDH) to 2.7 with the application of 10g LDH kg^?1 soil. It is concluded that the LDH has a potential to be used as a long-term nitrate exchanger to control the movement of nitrate in soil, and thereby reduce risks of nitrate leaching in crop production in sensible areas.     

A Nonparametric Test for Random Dropouts

The problem of dropout is a common one in longitudinal studies. One usually assumes for the analysis that dropout is at random. There are some tests to investigate this assumption. But these tests depend on normally distributed data or lack power, cf. Listing and Schlittgen (1998). We here propose an overall test which combines several Wilcoxon rank sum tests. The alternative hypothesis states that there is a tendency for larger (smaller) values of the target variable the last time the probands show up. The test is applicable with many ties also. It proves to perform well, compared to the test developed for normally distributed data, as well as to a test for completely missing at random which is proposed by Little (1988). An application to real data is given too.     

DNA Sequences of Frameshift and Other Mutations Induced by Icr-170 in Yeast

ICR-170-induced mutations in the CYC1 gene of the yeast Saccharomyces cerevisiae were investigated by genetic and DNA sequence analyses. Genetic analysis of 33 cyc1 mutations induced by ICR-170 and sequence analysis of eight representatives demonstrated that over one-third were frameshift mutations that occurred at one site corresponding to amino acid positions 29-30, whereas the remaining mutations were distributed more-or-less randomly, and a few of these were not frameshift mutations. The sequence results indicate that ICR-170 primarily induces G.C additions at sites containing monotonous runs of three G.C base pairs. However, some (Formula: see text) sites within the CYC1 gene were not mutated by ICR-170. Thus, ICR-170 is a relatively specific mutagen that preferentially acts on certain sites with monotonous runs of G.C base pairs.     

Synthesis of Apolipoprotein A-1 in Pig Brain Microvascular Endothelial Cells

In an approach toward the identification of hitherto unknown proteins involved in the function of the blood-brain barrier, we constructed a pig brain microvessel-derived cDNA library that is enriched in blood-brain barrier specific sequences by means of subtractive cloning. Sequence analysis of selected clones revealed that one of the cDNAs encoded porcine apolipoprotein (apo) A-1. The identity of apo A-1 mRNA was further confirmed by in vitro translation of RNA from brain microvascular endothelial cells and subsequent immunoprecipitation with an antibody against human apo A-1. We further investigated the expression of apo A-1 mRNA in several tissues and in endothelial cells of the pig. It is shown that cultured brain microvascular endothelial cells provide an in vitro model to study the expression and function of apo A-1 in the microvasculature of the brain.     

The Bacillus subtilis small cytoplasmic RNA gene and ''dnaX'' map near the chromosomal replication origin.

Summary TheBacillus subtilis small cytoplasmic RNA (scRNA) has an important, although not yet defined function in protein biosynthesis. Here we describe the mapping of the single copy scRNA gene and the flanking homolog todnaZX ofEscherichia coli, termed dnaX. The scRNA gene region of aB. subtilis wild-type strain was marked with acat gene and mapped by scoring chromosomal co-transformation rates of various mutant strains to chloramphenicol resistance and loss of the mutant phenotypes, respectively. This analysis, together with anEcoRI map comparison, places the scRNA gene anddnaX in the vicinity ofrecM near the replication origin region ofB. subtilis.     

Effects of high-frequency pressure waves applied to upper airway on respiration in central apnea.

We examined the effects of high-frequency (30-Hz) low-pressure oscillations on respiration in nine patients with central sleep apnea. All patients were studied during sleep and wore a nasal mask through which the oscillations were applied. All tests were performed during periods of repetitive central apneas. Respiratory efforts were monitored from the airflow and calibrated Respitrace signals. After several cycles of apnea were monitored, the oscillatory pressures were applied for brief periods (less than 5 s) at the midpoint of the central apneas. All trials in which arousal occurred were discarded, leaving a total of 106 trials in the nine patients. High-frequency oscillation of the upper airway stimulated respiratory effort(s) in 68% of all trials (72 of 106). Apnea length was significantly shortened in four of the nine patients. In one patient with a tracheostomy, the stimulus applied to his isolated upper airway evoked respiratory efforts during central apnea in 13 of 15 trials. We conclude that high-frequency oscillatory pressures applied to the upper airway can stimulate respiratory efforts during central apnea. This response may be mediated by upper airway receptors involved in nonrespiratory airway defense reflexes and may have implications in the treatment of patients with central sleep apnea.     

Stress ulcer modulation by limbic system structures.

A number of studies suggest that the telencephalic limbic system modulates stress ulcer development. The amygdala is assumed to connect sensory experiences, including stressful stimuli, with the emotional reactions and gastrointestinal effects normally produce. The hippocampal formation (entorhinal cortex, dentate gyrus, hippocampus) is part of a gating system, modulating the organism's coping ability. Changes in transmission in this temporal brain region are linked to individual differences in stress ulcer severity. Interactions among "classical" transmitters and several neuropeptides mediate these differences.