Midsummer heat wave effects on lacustrine plankton: Variation of assemblage structure and fatty acid composition

The effects of the 2003 European heat wave on a freshwater plankton assemblage and its fatty acid (FA) composition were investigated. Composition and FA profiles of four size categories of planktonic organisms collected in 2003 were compared to those of the colder year 2002.     

Separation of peptides by reverse-phase high-performance liquid chromatography using propyl- and cyanopropylsilyl supports

The separation of peptides and proteins by reverse-phase high-performance liquid chromatography with cyanopropylsilyl and large-pore propylsilyl supports, together with aqueous trifluoroacetic acid/acetonitrile gradients, was studied. Operating parameters (trifluoroacetic acid concentration, flow rate, and gradient slope) were evaluated using different enzymatic digests of horse cytochrome c and bovine serum albumin. Peptides ranging in size from five amino acids to 68 kDa could be separated on the propylsilyl column in a single chromatographic run. The cyanopropylsilyl column is suitable as a supplement to the use of the large-pore column for medium size (5-20 amino acids) peptides. The chromatographic supports and conditions presented here offer a simple, sensitive, and rapid separation system for a wide size range of peptides and proteins. They extend the versatility of separation methodology for these molecules.     

The Separation and Characterization of Carbohydrate Rich Component from Ovomucin in Chicken Eggs

We investigated the effects of near-infrared irradiation on the photoconversion of Chenopodium album water-soluble chlorophyll-binding protein (CaWSCP) in the presence of sodium hydrosulfite and found a further photoconversion from CP742 to CP763, a novel form of CaWSCP. Interestingly, one-third of the absorption peak at 668 nm was recovered in CP763, but re-irradiation under oxidative conditions eliminated the photo convertibility of CaWSCP.     

Quantitative assay of deoxyribonuclease activity after isoelectric focusing in polyacrylamide gels: pH control and effects of enzyme diffusion

A method for the quantitative assay of nuclease activity in crude cell lysates after isoelectric focusing (IEF) in polyacrylamide slab gels is described. After IEF, an agarose overlay gel containing DNA is placed on the IEF gel and the nuclease activity quantified by the loss of ethidium bromide fluorescence of the DNA. With this method a linear response was obtained for 1 to 10 ng of DNase I. Various methods of pH equilibration after IEF were also evaluated. The use of a high buffer concentration in the overlay gel is recommended to control the pH during the enzyme reaction. An analytical solution for the diffusion of enzymes from the IEF gel to the overlay gel is also presented and an equation that may be used to choose optimum times for transfer of the enzyme from the IEF gel to the overlay gel is given.     

Factors determining the reconstructive denaturation of proteins in sodium dodecyl sulfate solutions. Further circular dichroism studies on structural reorganization of seven proteins

The factors determining the onset and extent of reconstructive denaturation of proteins were considered by comparing circular dichroism (CD) data of seven proteins and previously published findings. The effects of sodium dodecyl sulfate (SDS) on the conformation of the following proteins were tested: lysozyme, the mitogens fromPhytolacca americana (fractions Pa2 and Pa4), lectin fromWistaria floribunda, ovine lutropin, a Bence Jones protein, and histone H2B. While the helix content of lysozyme was raised by SDS slightly, in the Bence Jones protein andW. floribunda lectin it increased from near zero to about 25–30%. In histone H2B the helix content was raised by SDS even to about 48%. However, no clear indication of helix formation could be observed in the mitogens and lutropin, even at low pH or 2.0–2.5. The tertiary structure of the proteins was perturbed by SDS. It was concluded that the reorganization of secondary structure of the proteins was favored by the following factors: (1) presence of helicogenic amino acid sequences in the protein, (2) availability of positively charged sites of the basic amino acids for interactions with the dodecyl ion, (3) absence of a large surplus of negatively charged sites on the surface of protein, and (4) absence of extensive disulfide cross-linking within the macromolecule. Both hydrophobic and electrostatic interactions occur in reconstructive denaturation, and the newly formed helices are stabilized by hydrophobic shielding by the alkyl chains of the alkyl sulfate.     

Pore-forming Activity of the Escherichia coli Type III Secretion System Protein EspD

Enterohemorrhagic Escherichia coli is a causative agent of gastrointestinal and diarrheal diseases. Pathogenesis associated with enterohemorrhagic E. coli involves direct delivery of virulence factors from the bacteria into epithelial cell cytosol via a syringe-like organelle known as the type III secretion system. The type III secretion system protein EspD is a critical factor required for formation of a translocation pore on the host cell membrane. Here, we show that recombinant EspD spontaneously integrates into large unilamellar vesicle (LUV) lipid bilayers; however, pore formation required incorporation of anionic phospholipids such as phosphatidylserine and an acidic pH. Leakage assays performed with fluorescent dextrans confirmed that EspD formed a structure with an inner diameter of ∼2.5 nm. Protease mapping indicated that the two transmembrane helical hairpin of EspD penetrated the lipid layer positioning the N- and C-terminal domains on the extralumenal surface of LUVs. Finally, a combination of glutaraldehyde cross-linking and rate zonal centrifugation suggested that EspD in LUV membranes forms an ∼280–320-kDa oligomeric structure consisting of ∼6–7 subunits.     

Current and emerging commercial optical biosensors.

The field of commercial optical biosensors is rapidly evolving, with new systems and detection methods being developed each year. This review outlines the currently available biosensor hardware and highlights unique features of each platform. Affinity-based biosensor technology, with its high sensitivity, wide versatility and high throughput, is playing a significant role in basic research, pharmaceutical development, and the food and environmental sciences. Likewise, the increasing popularity of biosensors is prompting manufacturers to develop new instrumentation for dedicated applications. We provide a preview of some of the emerging commercial systems that are dedicated to drug discovery, proteomics, clinical diagnostics and routine biomolecular interaction analysis.     

Photosynthetic pigment concentration, organization and interconversions in a pale green Syrian landrace of barley (Hordeum vulgare L., Tadmor) adapted to harsh climatic conditions

Tadmor is a Syrian barley landrace that has adapted to semi-arid environments. Its leaves are pale green because of a 30% decrease in the chlorophyll and the carotenoid content of the chloroplasts (leading to a 7·5% decrease in light absorption) compared with barley genotypes that are not adapted to harsh Mediterranean climatic conditions (e.g. Plaisant). This difference in pigment content was attenuated during growth of the plants in strong light, but was strongly amplified when strong light was combined with a high growth temperature. The low pigment content of Tadmor leaves was not associated with significant changes in the pigment distribution between the photosystems or between the reaction centres of the photosystems and their associated chlorophyll antennae. No significant difference in the photosynthetic activity (O₂ production per unit absorbed light) was observed between Tadmor and Plaisant. The conversion of violaxanthin to zeaxanthin in strong light and its reversal in darkness were much faster and operated at a higher capacity in Tadmor leaves compared with Plaisant leaves, resulting in an increased photostability of photosystem II in the former leaves. The accelerated xanthophylls interconversion in the Syrian landrace was associated with, and possibly related to, an increased fluidity of the thylakoid membranes. The lipid peroxide level was lower in Tadmor compared with Plaisant. In contrast, no difference was found in the non-photochemical quenching of chlorophyll fluorescence between the two barley genotypes. The data indicate that the pale green Syrian landrace is equipped to survive excessive irradiance through a passive reduction of the light absorptance of its leaves, which mitigates the heating effects of strong light, and through the active protection of its photochemical apparatus by a rapid xanthophyll cycling.     

1H, 13C and 15N resonance assignment of phage T4 endoribonuclease RegB

RegB is involved in the control of the phage T4 life cycle. It inactivates the phage early mRNAs when their translation is no more required. We determined its structure and identified residues involved in substrate binding. For this, all backbone and 90% of side-chain resonance frequencies were assigned.