Lipid-induced organization of a primary amphipathic peptide: a coupled AFM-monolayer study

To better understand the nature of the mechanism involved in the membrane uptake of a vector peptide, the interactions between dioleoylphosphatidylcholine and a primary amphipathic peptide containing a signal peptide associated with a nuclear localization sequence have been studied by isotherms analysis of mixed monolayers spread at the air-water interface. The peptide and the lipid interact through strong hydrophobic interactions with expansion of the mean molecular area that resulted from a lipid-induced modification of the organization of the peptide at the interface. In addition, a phase separation occurs for peptide molar fraction ranging from about 0.08 to 0.4 Atomic force microscopy observations made on transferred monolayers confirm the existence of phase separation and further reveal that mixed lipid-peptide particles are formed, the size and shape of which depend on the peptide molar fraction. At low peptide contents, round-shaped particles are observed and an increase of the peptide amount, simultaneously to the lipidic phase separation, induces morphological changes from bowls to filamentous particles. Fourier transform infrared spectra (FTIR) obtained on transferred monolayers indicate that the peptide adopts a beta-like structure for high peptide molar fractions. Such an approach involving complementary methods allows us to conclude that the lipid and the peptide have a nonideal miscibility and form mixed particles which phase separate.     

Batch treatment of liquid fraction of pig slurry by intermittent aeration: process simulation and microbial community analysis

BACKGROUND: Manure treatment in intensive livestock farming is required to reduce the risk of negative environmental impacts by nitrogen disposal. Biological removal through intermittent aeration in a single bioreactor is a suitable method for this purpose. The characteristic operation based on the alternation of oxic/anoxic phases confers these systems with certain particularities in terms of process modelling and of understanding the microbial interactions. RESULTS: The performance of a sequencing batch reactor (SBR) treating raw liquid fraction of pig slurry (LFPS) under loading rates of 0.13 g N L^?1 d^?1 was studied. Three different aeration strategies were applied: (1) constant airflow; (2) dissolved oxygen (DO) set‐point; and (3) DO‐based real‐time control. The comparatively low aeration intensity applied throughout the third strategy resulted in the process being performed mainly via nitrite, implying a reduction on the requirements of organic carbon and oxygen. However, a decrease in the nitrification rate was observed under those conditions. Experimental results were satisfactorily simulated by a mathematical model focused on organic carbon and nitrogen removal. Microbial community structure analysis through denaturing gradient gel electrophoretic profiling of 16S rDNA genes showed that the aeration exerted a strong influence on the dominant microbial populations. The presence of the ammonia‐oxidizing species Nitrosomonas europaea, and of denitrifying bacteria related to Thauera sp. and Ralstonia sp., was detected in the strategy at low DO. CONCLUSION: Dependence of model statement and parameter values on the bioreactor operational patterns and piggery wastewater composition was evidenced. Oxygen limitation was responsible for a significant microbial shift in SBR treating LFPS. Copyright © 2009 Society of Chemical Industry     

Viability, efficacy, and storage stability of freeze-dried biocontrol agent Candida sake using different protective and rehydration media

Viability, efficacy against Penicillium expansum on Golden Delicious apples, and storage stability of freeze-dried Candida sake strain CPA-1 were studied. The effect of several protective agents and rehydration media was investigated in the freeze drying of C. sake. Skimmed milk at 10% concentration was a good rehydration medium for all protectants tested. In general, good viability results were obtained when the same solution was used as a protectant and as a rehydration medium. The best survival was obtained when C. sake cells were protected with 10% lactose + 10% skimmed milk and rehydrated with skimmed milk (85% viability). The potential for biocontrol of the best freeze-dried treatments against P. expansum on apples was compared with that of fresh cells. Freeze-dried treatments at 1 x 10(7) CFU/ml reduced the incidence of decay by 45 to 66%. The best biocontrol effect was obtained with cells that had been freeze dried using 10% lactose + 10% skimmed milk as a protectant and 1% peptone as a rehydration medium, with a 66% reduction in rot incidence. However, in all treatments, the efficacy of freeze-dried cells was significantly lower than fresh cells. The stability of freeze-dried samples decreased during storage and was influenced by storage temperature. In the best treatment, storage of C. sake cells for 60 days at 4 degrees C resulte in final concentrations of 2.5 x 10(8) CFU/ml, which was a 10-fold reduction in relation to the initial starting concentration of cells prior to freeze drying.     

Effect of freeze drying and protectants on viability of the biocontrol yeast Candida sake

The effects of freezing method, freeze drying process, and the use of protective agents on the viability of the biocontrol yeast Candida sake were studied. Freezing at -20 degrees C was the best method to preserve the viability of C. sake cells after freeze drying using 10% skim milk as a protectant (28.9% survival). Liquid nitrogen freezing caused the highest level of damage to the cells with viability < 10%. Different concentrations of exogenous substances including sugars, polyols, polymers and nitrogen compounds were tested either alone or in combination with skim milk. There was little or no effect when additives were used at 1% concentration. Galactose, raffinose and sodium glutamate at 10% were the best protective agents tested alone but the viability of freeze-dried C. sake cells was always < 20%. Survival of yeast cells was increased from 0.2% to 30-40% by using appropriate protective media containing combinations of skim milk and other protectants such as 5% or 10% lactose or glucose, and 10% fructose or sucrose.     

Influence of co-winemaking technique in sensory characteristics of new Spanish red wines

The influence of co-winemaking technique on the sensory profile of wines made from minority red grape varieties cultivated in La Mancha region (Spain) was researched by sensory characterization. Sensory analysis was carried out by a panel of 15 experienced wine-testers, all with a previous training. The sensory profiles of monovarietal red wines—Cencibel, Bobal and Moravia Agria—were studied and also the effects of co-winemaking technique on sensory characteristics were evaluated. Co-winemaking red wines were obtained by blending (a) Cencibel (50%) + Bobal (50%); (b) Cencibel (50%) + Moravía Agria (50%) and (c) the three grape varieties Cencibel (33%) + Bobal (33%) + Moravía Agria (33%). The co-winemaking technique intensified the principal sensory properties of monovarietal wines and the wines proved to be more complex than the monovarietal wines.     

Dynamic extension - A new principle for a vacuum primary standard for UHV-range (10-10 Pa) calibration of hot cathode ionisation gauges

The main obstacle in the construction of primary vacuum standards for very low pressures - ultra-high vacuum (UHV) - is outgassing from the walls of the apparatus. Thus, principles utilising a higher pumping rate in the calibration chamber, e.g., the pressure ratio technique, seem to be more useful for this range. However, a serious problem is inherent in this method, namely, that the gas in the calibration chamber is not in equilibrium and thus cannot be characterised by a single scalar pressure value, although such a value is then used to evaluate the measurement. This discrepancy can be overcome by using a hot cathode ionisation gauge as a transfer standard calibrated by the dynamic extension method.The dynamic extension method is explained and compared with the pressure ratio technique. The basic conditions for achieving acceptable uncertainty values are considered.     

On the Influence of Polyvinylpyrrolidone (PVP) during the Synthesis of Ultrafine Emulsions

The influence of PVP as well as the influence of the excess of silver ions, and the quantity of silver iodide in the crystal phase, on the physical ripening of Lipmann’s emulsions are investigated. It is established that PYP favours the formation of ultrafine emulsions. Its inhibitory influence on physical ripening makes it possible to regulate and equalize the activity of the gelatins, used in the synthesis of Lipmann’s emulsions.

Emulsions with a resolving power of 3200 1/mm satisfactory sensitivity and edge sharpness are synthesized.     

Purification, molecular cloning, and functional expression of the human nicodinamide-adenine dinucleotide phosphate-regulated thyroid hormone-binding protein

The kidney and several other thyroid hormone-responsive tissues contain a NADP-regulated thyroid hormone (TH)-binding protein (THBP), with an apparent molecular mass of 36 kDa on SDS-PAGE, responsible for most of the intracellular high-affinity T3 and T4 binding. THBP was purified to homogeneity from human kidney cytosol and used to generate proteolytic peptides. Microsequencing of four peptides revealed identity to amino acid sequences deduced from a human cDNA homolog to a cDNA encoding kangaroo mu-crystallin. This protein is a major structural kangaroo lens protein with no known function in other species. A full-sized cDNA (TH5.9) was isolated by 5'- and 3'-rapid amplification of cDNA ends using a human brain cDNA library and gene-specific PCR primers, confirming identity to the previously cloned human cDNA. The TH5.9 cDNA encodes a 314-residue protein (theoretical mol wt = 33,775) with significant homologies (40 to 60%) with two bacterial enzymes: lysine cyclodeaminase and ornithine cyclodeaminase. The TH5.9 cDNA was expressed in Escherichia coli as a glutathione S-transferase (GST) fusion protein. Purified GST fusion protein, but not GST, bound T3 specifically with high affinity [dissociation constant (Kd) = 0.5 nM] in the presence of NADPH, and was labeled by UV-driven cross-linking of underivatized [(125)I]T3. T3 binding and photoaffinity labeling of GST fusion protein were activated by NADPH [activation constant (K[act]) = 10(-8) M], but not by NADH. The expressed protein displays the appropriate binding properties, indicating that TH5.9 cDNA encodes the NADP-regulated THBP characterized in human tissues.