Sodium-dependent uptake of taurine in encapsulated Staphylococcus aureus strain M

A novel uptake system for the unusual sulfonated amino acid taurine was discovered in the prokaryote, encapsulated Staphylococcus aureus strain M. This strain has been shown previously to contain taurine in its capsular polysaccharide. Taurine uptake by whole cells incubated in buffer showed a saturable dependency upon Na⁺ and taurine uptake was itself a saturable process, stimulated by glucose, and markedly affected by temperature. No evidence was found for the inducibility of taurine uptake. In the presence of 10 mM NaCl Lineweaver-Burk plots revealed a K_m of 42 μM and V_max of 4.6 nmol/min per mg dry weight for taurine uptake at 37°C. Increasing concentrations of Na⁺ decreased the K_m of the system and appeared to increase the V_max. Of various other cations tested only Li⁺ supported marked taurine uptake. Excess unlabelled taurine did not cause efflux of radioactivity taken up. Taurine was taken up into cold trichloroacetic acid-soluble material and did not chromatograph as taurine, indicating rapid metabolism during or closely following uptake. Taurine uptake appeared to occur via a highly specific system because amino acids representing the major known groups of amino acid transport systems in S. aureus did not inhibit taurine uptake, and uptake was only slightly diminished by the structurally closely related compounds hypotaurine and 3-amino-1-propane sulfonic acid. Sulfhydryl group reagents, electron transport inhibitors, an uncoupler and inhibitors of Na⁺-linked transport processes inhibited taurine uptake. A variety of other metabolic inhibitors had little effect on taurine uptake.     

SilE is an intrinsically disordered periplasmic “molecular sponge” involved in bacterial silver resistance

Ag⁺ resistance was initially found on the Salmonella enetrica serovar Typhimurium multi‐resistance plasmid pMG101 from burns patients in 1975. The putative model of Ag⁺ resistance, encoded by the sil operon from pMG101, involves export of Ag⁺ via an ATPase (SilP), an effluxer complex (SilCFBA) and a periplasmic chaperon of Ag⁺ (SilE). SilE is predicted to be intrinsically disordered. We tested this hypothesis using structural and biophysical studies and show that SilE is an intrinsically disordered protein in its free apo‐form but folds to a compact structure upon optimal binding to six Ag⁺ ions in its holo‐form. Sequence analyses and site‐directed mutagenesis established the importance of histidine and methionine containing motifs for Ag⁺‐binding, and identified a nucleation core that initiates Ag⁺‐mediated folding of SilE. We conclude that SilE is a molecular sponge for absorbing metal ions.     

α-Factor inhibition of the rate of cell passage through the “start” step of cell division in Saccharomyces cerevisiae yeast: Estimation of the division delay per α-factor·receptor complex

A highly sensitive, kinetically unambiguous assay for α-factor-induced delay of cell passage through the “start” step of cell division in yeast is presented. The assay employs perfusion with periodic microscopy to monitor the bud emergence kinetics on the 20% of cells within an exponentially growing population which exist prior to the α-factor execution point of start. The t_1/2 for cell passage through start by this population of cells is 31 min in the absence of α-factor. The inhibition constant, K_I, represents the α-factor concentration which produces a 50% inhibition of this rate and is equal to 2×10⁻¹⁰M. A second assay for maximal cell division arrest by α-factor on whole populations of cells is presented. This assay shows a maximum cell division arrest time of 125±5 h at saturating α-factor, and a K₅₀ (that is, an α-factor concentration which produces a half-maximal response) of 2.5×10⁻⁸M. Both assays were performed in the effective absence of α-factor inactivation. Values of the dissociation constant K_D and total number of receptors per cell which specifically mediate cell division arrest or delay were estimated to be 2.5×10⁻⁸M and 10⁴, respectively. These estimates, along with the quantitative dose-response data for division arrest which are presented here, are consistent with each receptor·α-factor complex which is present on the cell at equilibrium producing a 43±10 s delay of cell passage through start. Surprisingly, this number is constant within twofold over the entire range of cellular division arrest responses to α-factor, that is, from a 1.9-fold inhibition of the rate of cell passage through start at 0.17 nM α-factor to a 125±5 h maximum arrest at saturating α-factor concentrations of >170 nM. The possible significance of this observation toward the mechanism of α-factor-induced cell division arrest is discussed.     

Carrier Transport and Recombination in Efficient “All‐Small‐Molecule” Solar Cells with the Nonfullerene Acceptor IDTBR

Reaching device efficiencies that can rival those of polymer‐fullerene Bulk Heterojunction (BHJ) solar cells (>10%) remains challenging with the “All‐Small‐Molecule” (All‐SM) approach, in part because of (i) the morphological limitations that prevail in the absence of polymer and (ii) the difficulty to raise and balance out carrier mobilities across the active layer. In this report, the authors show that blends of the SM donor DR3TBDTT (DR3) and the nonfullerene SM acceptor O‐IDTBR are conducive to “All‐SM” BHJ solar cells with high open‐circuit voltages (V_OC) >1.1 V and PCEs as high as 6.4% (avg. 6.1%) when the active layers are subjected to a post‐processing solvent vapor‐annealing (SVA) step with dimethyl disulfide (DMDS). Combining electron energy loss spectroscopy (EELS) analyses and systematic carrier recombination examinations, the authors show that SVA treatments with DMDS play a determining role in improving charge transport and reducing non‐geminate recombination for the DR3:O‐IDTBR system. Correlating the experimental results and device simulations, it is found that substantially higher BHJ solar cell efficiencies of >12% can be achieved if the IQE and carrier mobilities of the active layer are increased to >85% and >10^?4 cm² V^?1 s^?1, respectively, while suppressing the recombination rate constant k to <10^?12 cm³ s^?1.     

Development of Bifidobacterium lactis Bb 12 on β‐(2,6)‐Linked Fructan‐Containing Substrate

β‐(2,1)‐linked fructan of plant origin (inulin) and the related oligosaccharides (FOS) as non‐digestible carbohydrates, i.e., potent prebiotics, can stimulate the growth of various probiotic lactic acid bacteria, including a number of bifidobacteria strains. The related β‐(2,6)‐linked fructans of microbial origin (levan and FOS), however, have scarcely been investigated in this respect. Therefore, the bifidogenic properties of various fructans, i.e., inulin, levan, fructooligosaccharides (FOS) and fructan syrup (FS), were tested as glucose substitutes in MRS media and were compared concerning their effect on the commercial strain Bifidobacterium lactis Bb 12. Although glucose was the preferred substrate for growth and biomass formation, FS exhibited a comparable cell growth (8.4 × 10⁷ counts/mL and 1.0 × 10⁷ counts/mL, respectively) and acidification power (84 °T and 74 °T, respectively) during 48 h of fermentation, as well as an increase in lactic acid and decrease in acetic acid formation. Bifidobacterium lactis Bb 12 did not utilize inulin as a sole carbon source as judged from the 60 % decrease in cell count and the insignificant (0.1 pH unit) acidification of the growth medium, whereas levan provided a noticeable increase in cell count and acidification (0.4 pH units) during 48 h of fermentation. FOS preparation appeared to be a satisfactory carbon source for this strain, but lower acidification power (56 °T) and cell counts were observed as compared to glucose‐ or FOS‐containing media (2.6 % and 22 %, respectively). The products obtained under conditions of mild lactic acid hydrolysis of levan (37 °C, pH 3.3, 24 h) enhanced the cell count (7–10 %) and acidification power (by a factor of 2.7) of Bifidobacterium lactis Bb 12.     

Molecular Characterization and In Situ Localization of a Mouse Retinal Taurine Transporter

Abstract: Various ocular tissues have a higher concentration of taurine than plasma. This taurine concentration gradient across the cell membrane is maintained by a high-affinity taurine transporter. To understand the physiological role of the taurine transporter in the retina, we cloned a taurine transporter encoding cDNA from a mouse retinal library, determined its biochemical and pharmacological properties, and identified the specific cellular sites expressing the taurine transporter mRNA. The deduced protein sequence of the mouse retinal taurine transporter (mTAUT) revealed >93% sequence identity to the canine kidney, rat brain, mouse brain, and human placental taurine transporters. Our data suggest that the mTAUT and the mouse brain taurine transporter may be variants of one another. The mTAUT synthetic RNA induced Na⁺- and Cl^?-dependent [³H]taurine transport activity in Xenopus laevis oocytes that saturated with an average K_m of 13.2 µM for taurine. Unlike the previous studies, we determined the rate of taurine uptake as the external concentration of Cl^? was varied, a single saturation process with an average apparent equilibrium constant (K_Cl?) of 17.7 mM. In contrast, the rate of taurine uptake showed a sigmoidal dependence when the external concentration of Na⁺ was varied (apparent equilibrium constant, K_Na+～54.8 mM). Analyses of the Na⁺- and Cl^?-concentration dependence data suggest that at least two Na⁺ and one Cl^? are required to transport one taurine molecule via the taurine transporter. Varying the pH of the transport buffer also affected the rate of taurine uptake; the rate showed a minimum between pH 6.0 and 6.5 and a maximum between pH 7.5 and 8.0. The taurine transport was inhibited by various inhibitors tested with the following order of potency: hypotaurine > β-alanine > l -diaminopropionic acid > guanidinoethane sulfonate > β-guanidinopropionic acid > chloroquine > γ-aminobutyric acid > 3-amino-1-propanesulfonic acid (homotaurine). Furthermore, the mTAUT activity was not inhibited by the inactive phorbol ester 4α-phorbol 12,13-didecanoate but was inhibited significantly by the active phorbol ester phorbol 12-myristate 13-acetate, which was both concentration and time dependent. The cellular sites expressing the taurine transporter mRNA in the mouse eye, as determined by in situ hybridization technique, showed low levels of expression in many of the ocular tissues, specifically the retina and the retinal pigment epithelium. Unexpectedly, the highest expression levels of taurine transporter mRNA were found instead in the ciliary body of the mouse eye.     

The effect of hyperpolarization of cell R15 on the hemolymph composition of intactAplysia

Summary Experiments were performed to determine the effects of the activity pattern in identified neurosecretory cell R15 on the distribution of osmolytes inAplysia californica. We have used an intact preparation which allows intracellular microelectrode penetration of R15 while circulatory and peripheral sensory inputs remain relatively undisturbed. Exposure of the animal to dilute sea water produces a reduction of bursting pacemaker activity in R15. The magnitude of burst frequency reduction is correlated with the degree to which the salinity is reduced. When R15 is silenced by passage of current through a microelectrode, a number of changes occur in the concentration of osmolytes in the hemolymph. These changes include a 13% increase in K⁺, and a 70% increase in ammonia levels. Hemolymph Na⁺ levels were unchanged by R15 hyperpolarization. The distribution of free amino acids was also affected by R15 hyperpolarization. The changes were selective, with the most pronounced effect being an increase of taurine levels of 367%. The role which R15 may play in osmoregulation is discussed in light of these findings.Abbreviations ASW artificial sea water - BPA bursting pacemaker activity - TCA trichloroacetic acid - FAA free amino acids     

Hydrology of winter–spring “red tides” in Bahía de Mazatlán, Sinaloa, México

“Red tide” events are frequent and periodical in Bahía de Mazatlán, Sinaloa, México. Yet, the ones observed from 4 February to 4 June 2000, showed some distinctive features: First, the dinoflagellates Prorocentrum balticum (85%), P. mexicanum (5%), and Ceratium furca (5%), dominated the composition of the blooms; Second, the average cell abundance by date was 1.3×10⁶ cells l⁻¹, with a range of 3.5×10³ to 24,500 × 10³ cells l⁻¹, well above previous records; Third, the temperature registered at 10–20 m deep was unusually cold (19 °C), below the normal average of 21.5 °C observed over the last 10 years. Salinity was high (35.9 psu) and showed very little influence on the water density gradient. A mean thermal stratification index (TSI), of 3.4, with a maximum of 7 °C, was observed throughout the period, in spite of a weak upwelling activity and intermittent strong NW winds which were unable to break up water column stratification. Temperature fluctuations at the surface and at the bottom layers showed a 30-day periodicity, suggesting some association with the lunar cycle. To explain the characteristics of the “red tides” registered in Bahía de Mazatlán during the winter–spring period of year 2000, it is proposed that the temperature and density stratification, stabilized further by internal waves that compensated for the weak upwelling activity and provided the necessary nutrients to the surface layer, favored the persistence and intensity of the harmful algal bloom events then observed.     

Rapid detection of “Candidatus Phytoplasma mali” by recombinase polymerase amplification assays

Isothermal recombinase polymerase amplification (RPA) assays for the specific detection of “Candidatus Phytoplasma mali (Ca. P. mali),” the causal agent of apple proliferation, were developed. The assays amplify a fragment of the imp gene and amplimers were detected either by fluorescence in real‐time mode (TwistAmp^®exo assay) using a fluorophore‐labelled probe or by direct visualization employing a lateral flow device (TwistAmp^®nfo assay/Milenia^®HybriDetect). The RPA assays specifically amplified DNA from “Ca. P. mali” strains, and cross‐reactivity with other phytoplasmas or plant DNA was not observed. The limit of detection was determined with a cloned imp standard, and positive results were obtained down to 10 copies with both RPA assay formats. In comparison with a TaqMan real‐time PCR assay based on the same target gene, the RPA assays were equally sensitive, but results were obtained faster. Simplified nucleic acid extraction procedures from plant tissue with Tris‐ and CTAB‐based buffers revealed that crude Tris–DNA extracts were a suitable source for RPA tests while larger concentrations of CTAB were inhibitory. This is the first report of RPA‐based assays for the detection of “Ca. P. mali”. The assays are suitable for high‐throughput screening of plant material and point‐of‐care diagnostic and can be potentially combined with a simplified DNA extraction procedure.     

Expression and Purification of a Recombinant “Small” Sialidase fromClostridium perfringensA99

A 1.4-kb gene encoding the “small” sialidase isoenzyme ofClostridium perfringensA99, including its own promoter, was previously cloned in and expressed byEscherichia coliJM 101. Since all attempts to purify this enzyme to homogeneity were unsuccessful, a new strategy was developed. The structural gene was amplified by means of a PCR technique and inserted into the plasmid vector pQE-10, transferring a six-histidine affinity tag (His₆) to the N-terminus of the protein. In order to minimize proteolytic degradation of the sialidase protein, the gene was subcloned into theEscherichia colistrain BL21(DE3)pLys S with reduced protease activity. The sialidase production was increased about 2.5-fold when compared with that of the original clone. The enzyme, released by lysozyme treatment of the bacterial cells, was purified by metal chelate chromatography on Ni–nitrilo-triacetic acid agarose to apparent homogeneity in SDS–PAGE. The 42-kDa protein was enriched 62-fold with a yield of 82% and a specific activity of 280 U mg⁻¹. A total amount of 1 mg sialidase was obtained from 1 liter of bacterial culture. For future studies, including crystallization experiments, the histidine affinity tag was removed from the sialidase enzyme by aminopeptidase K. The sialidase was then separated from aminopeptidase K by ion-exchange chromatography, resulting in an overall yield of 83% and a specific activity of 305 U mg⁻¹using 4-methylumbelliferyl-α- -N-acetylneuraminic acid under standard conditions. The two forms (with or without the histidine tag) of sialidase exhibited similar kinetic properties when compared to the wild-type enzyme.     

“Ecological Armageddon” – more evidence for the drastic decline in insect numbers

The efficient use of nitrogen by crops can minimise environmental risks and maximise returns to farmers. Under organic farming systems, this can be achieved by adjusting the fertilisation management and/or using genetic variability. Seven durum wheat (Triticum durum) cultivars and three emmer (Triticum dicoccum) cultivars were assessed under an organic farming system over a non‐consecutive 4‐year period (2005–11) in Foggia (southern Italy). The objectives were to investigate the agronomic and qualitative characteristics, and to evaluate the agronomic efficiency and adaptability according to three N fertilisation levels (0, 40, 80 kg N ha^?1). A split‐plot design was used, with three replications in each year. Nine traits were investigated: heading time, plant height, seed yield, number of spikes m^?2, harvest index, specific weight, 1000‐seed weight, and protein and gluten contents. Increasing N to 80 kg ha^?1 increased plant height (+7.3%), seed yield (+22.1%), spike density (+16.6%), and protein (+9.8%) and gluten (+2.1%) contents. The emmer genotypes and the oldest durum wheat ‘Cappelli’ showed the highest protein and gluten contents (mean, 13.9%, 11.2%, respectively). The maximum N agronomic efficiency [AE(N)] and N recovery efficiency [RE(N)] were seen for the modern durum wheat cultivars under 40 kg ha^?1 N treatment: ‘Duilio’, ‘Iride’ and ‘Varano’. The significant correlation between AE(N) and RE(N) and the year of release of the durum wheat cultivars (r = 0.53, P < 0.05, F = 4.7 for AE(N) and r = 0.57, P < 0.01, F = 5.7 for RE(N), respectively; n = 14) showed that the genetic breeding of this species for high grain yield is associated with an increase in the efficiency of N fertiliser use under organic farming. The highly significant effect for the Genotype × Environment interaction (F = 18.1, P < 0.001) of seed yield indicate the possibility to select for stable genotypes across environments. Based on the additive main effects and multiplicative interaction analysis, and the yield stability index, the varieties ‘Iride’ and ‘Varano’ show high‐stability responses and good seed yields under all N fertiliser treatments. Therefore, these varieties can be recommended for organic farming systems in Mediterranean areas.     

Modified nitrocefin‐EDTA method to differentially quantify the induced L1 and L2 β‐lactamases in Stenotrophomonas maltophilia

Aim: To estimate the ethylene diamine tetraacetic acid (EDTA) concentration at which the L1 enzyme activity in the cell extracts of Stenotrophomonas maltophilia can be mostly inhibited. Methods and Results: The effective inhibition concentration of EDTA against the L1 enzyme in the cell extracts was firstly evaluated by using the L2 isogenic mutant of S. maltophilia KJ, KJΔL2, as the assayed strain. Approximately 92% L1 activity was inhibited by 10 mmol l^?1 EDTA, which is 100‐fold higher than that from previously reported protocols (0·1 mmol l^?1). Three phylogenetic clusters of L1 proteins were revealed from 11 clinical S. maltophilia isolates, with a L1 protein divergence of 0–11%. The EDTA concentration required to inhibit the L1 enzymes of different phylogenetic clusters was estimated to be 10 mmol l^?1. Conclusion: The previous nitrocefin‐EDTA protocol for differentially quantifying the L1 and L2 activity in the cell extracts has been modified by raising the added EDTA concentration to 10 mmol l^?1. Significance and Impact of the Study: A rapid and accurate method for determination of L1 and L2 activity will provide a convenient tool for enzyme characterization and induction mechanism study of S. maltophilia.     

Parameters of unbalanced growth and reversible inhibition of deoxyribonucleic acid synthesis in Brevibacterium ammoniagenes ATCC 6872 induced by depletion of Mn2+. Inhibitor studies on the reversibility of deoxyribonucleic acid synthesis

Unbalanced growth induced by depletion of manganese ions was a prerequisite for production of ribonucleotides in a high salt mineral medium with the wildtype strain Brevibacterium ammoniagenes ATCC 6872. The concentration of manganese strictly controlled the overall deoxyribonucleic acid (DNA) synthesis, whereas ribonucleic acid (RNA), protein and cell wall synthesis remained essentially unimpaired in the manganese-lacking cells.The reversibility of inhibition of overall DNA synthesis was shown by enhanced incorporation (up to threefold compared to the cultures supplied with sufficient manganese) of [8-¹⁴C] adenine into alkali-stable, trichloroacetic acid-insoluble material after subsequent addition of 10 M MnCl₂ to 15 h-old depleted cultures.The results of inhibitor studies on the restoration of overall DNA synthesis due to subsequent addition of manganese ions to depleted cultures suggest that ribonucleotide reduction is the primary target of the manganese starvation during nucleotide fermentation with Brevibacterium ammoniagenes ATCC 6872.     

Dietary Restraint and Control Over “Wanting” Following Consumption of “Forbidden” Food

Eating behavior can be influenced by the rewarding value of food, i.e., “liking” and “wanting.” The objective of this study was to assess in normal‐weight dietary restrained (NR) vs. unrestrained (NU) eaters how rewarding value of food is affected by satiety, and by eating a nonhealthy perceived, dessert‐specific food vs. a healthy perceived, neutral food (chocolate mousse vs. cottage cheese). Subjects (24NR age = 25.0 ± 8.2 years, BMI = 22.3 ± 2.1 kg/m²; 26NU age = 24.8 ± 8.0 years, BMI = 22.1 ± 1.7 kg/m²) came to the university twice, fasted (randomized crossover design). Per test‐session “liking” and “wanting” for 72 items divided in six categories (bread, filling, drinks, dessert, sweets, stationery (placebo)) was measured, before and after consumption of chocolate mousse/cottage cheese, matched for energy content (5.6 kJ/g) and individual daily energy requirements (10%). Chocolate mousse was liked more than cottage cheese (P < 0.05). After consumption of chocolate mousse or cottage cheese, appetite and “liking” vs. placebo were decreased in NR and NU (P < 0.03), whereas “wanting” was only decreased in NR vs. NU (P ≤ 0.01). In NR vs. NU “wanting” was specifically decreased after chocolate mousse vs. cottage cheese; this decrease concerned especially “wanting” for bread and filling (P < 0.05). To conclude, despite similar decreases in appetite and “liking” after a meal in NR and NU, NR decrease “wanting” in contrast to NU. NR decrease “wanting” specifically for a nonhealthy perceived, “delicious,” dessert‐specific food vs. a nutritional identical, yet healthy perceived, slightly less “delicious,” “neutral” food. A healthy perceived food may thus impose greater risk for control of energy intake in NR.     

Enantiomeric assay of escitalopram S(+)‐enantiomer and its “in‐process impurities” using two different techniques

The enantiomeric purity of escitalopram oxalate ESC and its “in‐process impurities,” namely, ESC‐N‐oxide, ESC‐citadiol, and R(?)‐enantiomer were studied in drug substance and products using high‐performance liquid chromatography (HPLC)‐UV (Method I), synchronous fluorescence spectroscopy (SFS) (Method IIA), and first derivative SFS (Method IIB). Method I describes as an isocratic HPLC‐UV for the direct resolution and determination of enantiomeric purity of ESC and its “in‐process impurities.” The proposed method involved the use of α_l‐acid glycoprotein (AGP) chiral stationary phase. The regression plots revealed good linear relationships of concentration range of 0.25 to 100 and 0.25 to 10 μg mL^?1 for ESC and its impurities. The limits of detection and quantifications for ESC were 0.075 and 0.235 μg mL^?1, respectively. Method II involves the significant enhancement of the fluorescence intensities of ESC and its impurities through inclusion complexes formation with hydroxyl propyl‐β‐cyclodextrin as a chiral selector in Micliavain buffer. Method IIA describes SFS technique for assay of ESC at 225 nm in presence of its impurities: R(?)‐enantiomer, citadiol, and N‐oxide at ?λ of 100 nm. This method was extended to (Method IIB) to apply first derivative SFS for the simultaneous determination of ESC at 236 nm and its impurities: the R(?)‐enantiomer, citadiol, and N‐oxide at 308, 275, and 280 nm, respectively. Linearity ranges were found to be 0.01 to 1.0 μg mL^?1 for ESC and its impurities with lower detection and quantification limits of 0.033/0.011 and 0.038/0.013 μg mL^?1 for SFS and first derivative synchronous fluorescence spectra (FDSFS), respectively. The methods were used to investigate the enantiomeric purity of escitalopram.     

Dopamine for “Wanting” and Opioids for “Liking”: A Comparison of Obese Adults With and Without Binge Eating

Obesity research suffers from an overinclusion paradigm whereby all participants with a BMI beyond a certain cutoff value (e.g., 30) are typically combined in a single group and compared to those of normal weight. There has been little attempt to identify meaningful subgroups defined by their salient biobehavioral differences. In order to address this limitation, we examined genetic and psychological indicators of hedonic eating in obese adults with (n = 66) and without (n = 70) binge eating disorder (BED). Our analyses focused on dopamine (DA) and opioid genetic markers because of their conjoint association with the functioning of brain reward mechanisms. We targeted three functional polymorphisms related to the D2 receptor (DRD2) gene, as well as the functional A118G polymorphism of the mu‐opioid receptor (OPRM1) gene. We found that significantly more obese controls had the “loss‐of‐function” A1 allele of Taq1A compared to their BED counterparts, whereas the “gain‐of‐function” G allele of A118G occurred with greater frequency in the BED group. A significant gene–gene combination χ² analysis also indicated that of those participants with the gain‐gain genotype (G⁺ and A1), 80% were in the BED group whereas only 35% with the loss‐loss genotype (G^? and A1⁺) were in this group. Finally, BED subjects had significantly higher scores on a self‐report measure of hedonic eating. Our findings suggest that BED is a biologically based subtype of obesity and that the proneness to binge eating may be influenced by a hyper‐reactivity to the hedonic properties of food—a predisposition that is easily exploited in our current environment with its highly visible and easily accessible surfeit of sweet and fatty foods.     

Metal dependent hydrolysis of β‐casein by sIgA antibodies from human milk

We present the first evidence that electrophoretically and immunologically homogeneous sIgAs purified from milk of healthy human mothers by chromatography on Protein A‐Sepharose and FPLC gel filtration contain intrinsically bound metal ions (Ca > Mg ≥ Al > Fe ≈ Zn ≥ Ni ≥ Cu ≥ Mn), the removal of which by a dialysis against ethylenediamine tetraacetic acid (EDTA) leads to a significant decrease in the β‐casein‐hydrolyzing activity of these antibodies (Abs). An affinity chromatography of total sIgAs on benzamidine‐Sepharose interacting with canonical serine proteases separates a small metalloprotease sIgA fraction (6.8 ± 2.4%) from the main part of these Abs with a serine protease‐like β‐casein‐hydrolyzing activity. The relative activity of this metalloprotease sIgA fraction containing intrinsically bound metal ions increases ～1.2–1.9‐fold after addition of external metal ions (Mg²⁺ > Fe²⁺ > Cu²⁺ ≥ Ca²⁺ ≥ Mn²⁺) but decreases by 85 ± 7% after the removal of the intrinsically bound metals. The metalloprotease sIgA fraction free of intrinsic metal ions demonstrates a high β‐casein‐hydrolyzing activity in the presence of individual external metal ions (Fe²⁺ > Ca²⁺ > Co²⁺ ≥ Ni²⁺) and especially several combinations of metals: Co²⁺ + Ca²⁺ < Mg²⁺ + Ca²⁺ < Ca²⁺ + Zn²⁺ < Fe²⁺ + Zn²⁺ < Fe²⁺ + Co²⁺ < Fe²⁺ + Ca²⁺. The patterns of hydrolysis of a 22‐mer oligopeptide corresponding to one of sIgA‐dependent specific cleavage sites in β‐casein depend significantly on the metal used. Metal‐dependent sIgAs demonstrate an extreme diversity in their affinity for casein‐Sepharose and chelating Sepharose, and interact with Sepharoses bearing immobilized monoclonal mouse IgGs against λ‐ and κ‐type light chains of human Abs. Possible ways of the production of metalloprotease abzymes (Abz) by human immune system are discussed. Copyright © 2010 John Wiley & Sons, Ltd.     

Cellular uptake of taurine by lactating porcine mammary tissue

Milk taurine plays a critical role in neonatal development. Taurine uptake in lactating sow mammary tissue has not been characterized previously. The kinetic properties, ion dependence and substrate specificity of taurine uptake were characterized in mammary tissue collected from lactating sows at slaughter. Tissue explants were incubated in an isosmotic physiologic buffer with [³H]taurine tracer to measure taurine uptake. Taurine uptake was dependent upon the presence of extracellular sodium and chloride ions, which is consistent with the co-transport of sodium and chloride with taurine. Uptake was not dependent upon ion exchange mechanisms or upon furosemide-sensitive ion co-transport. Taurine uptake was saturable and exhibited an apparent K_m of 20 μM and a V_max of 386 μmol/kg cell water/30 min. Substrate specificity studies indicated a strong interaction of β-amino acids with the taurine transport system. Taurine transport in lactating sow mammary tissue is therefore a high affinity, sodium-dependent mechanism specific for β-amino acids, and is analogous to sodium-dependent taurine uptake in other tissues. The high affinity and high specificity of the taurine uptake system allows for concentration of taurine within the mammary cell and is ultimately responsible for provision of taurine required for neonatal development.     

Interaction of β‐cyclodextrin‐capped CdSe quantum dots with inorganic anions and cations

A facile method was developed for the preparation of water soluble β‐Cyclodextrin (β‐CD)‐modified CdSe quantum dots (QDs) (β‐CD‐QDs) by directly replacing the oleic acid ligands on the QDs surface with β‐CD in an alkaline aqueous solution. The as‐prepared QDs show good stability in aqueous solution for several months. Oxoanions, including phosphoric acid ion, sulphite acid ion and carbonic acid ion, affect the fluorescence of β‐CD‐QDs. Among them, H₂PO₄^– exhibited the largest quenching effect. For the polyprotic acids (HO)₃AO, the effect of acidic anions on the fluorescence of β‐CD‐QDs was in the order: monoanion (HO)₂AO₂^– > dianion (HO)AO₃^2– >> trianion AO₄^3–. After photoactivation for several days in the presence of anions at alkaline pH, the β‐CD‐QDs exhibited strong fluorescence emission. The effect of various heavy and transition metal ions on the fluorescence properties of the β‐CD‐QDs was investigated further. It was found that Ag⁺, Hg²⁺ and Co²⁺ have significant quenching effect on the fluorescence of the β‐CD‐QDs. The Stern–Volmer quenching constants increased in the order: Hg²⁺ < Co²⁺ <Ag⁺. The adsorption model of metal ions on β‐CD‐QDs was explored. Copyright © 2011 John Wiley & Sons, Ltd.