Chimeras of the homing endonuclease PI-SceI and the homologous Candida tropicalis intein: a study to explore the possibility of exchanging DNA-binding modules to obtain highly specific endonucleases with altered specificity

Homing endonucleases are extremely specific endodeoxyribonucleases. In vivo, these enzymes confer mobility on their genes by inducing a very specific double-strand cut in cognate alleles that lack the cooling sequence for the homing endonuclease; the cellular repair of the double-strand break with the endonuclease-containing allele as a template leads to integration of the endonuclease gene, completing the homing process. As a result of their extreme sequence specificity, homing endonucleases are promising tools for genome engineering. For this purpose, it is desirable to design enzymes with defined new specificities. To analyse which DNA-binding elements are potential candidates for use in the design of enzymes with modified or even new specificity, we produced several chimeric proteins derived from the Saccharomyces cerevisiae VMA1 intein (PI-SceI) and the related Candida tropicalis VMA1 intein. Although the mature Candida intein is devoid of endonucleolytic activity, the exchange of two DNA-binding modules of PI-SceI with the homologous elements from the Candida intein results in an active endonuclease. The low sequence homology in these modules indicates that different protein-DNA contacts are responsible for the recognition of related DNA sequences. This flexibility in DNA recognition should, in principle, allow endonucleases to be produced with new specificities useful for genome engineering.     

Nitrogen and phosphorus uptake,yield and agronomic traits of oat cultivars as affected by fertilizer N rates under diverse environments

Oat has been gaining renewed interest due to its role in human healthy diet. A field study was conducted across three diverse locations in Canada to determine N and P uptake, agronomic traits and yield performance of 10–12 cultivars under four fertilizer N rates. Our data showed that ‘SA060123’ and ‘OA1331-5’ were the highest-yielding cultivars, and ‘Dieter’ and ‘Morrison’ the lowest, across sites-years. Yield components were altered to adapt to soil-environmental conditions, specifically, panicles m^?2 mostly accounted for yield variation at Melfort, seeds panicle^?1 and 1000-seed weight at Normandin, and lodging index was an additional yield-determining factor at Ottawa. It was noted that severe crop lodging that occurred mainly at Ottawa, was logically associated with greater accumulation of straw N and plant biomass under high N supply conditions, and that crop lodging displayed a strong correlation with straw P content. Relationship of lodging index and straw P was best-fit into a split-line model with a change-point of 13.6 kg P ha^?1, below which lodging rarely occurred. We speculate that high straw P content, induced by external N supply, may have exhibited similar behavior as N in weakening the strength of stem base and anchorage system, leading to crop lodging. This study also demonstrated an interactive genotype-by-environment effect on all traits except for the seed number panicle^?1. Multidimensional preference analysis revealed the best performance of ‘SA060123’ for outstanding yield and 'Minstrel' for harvest index, respectively.     

Effect of increase of dietary polyunsaturated fatty acids (omega 6) on the hepatic glutathione content and gamma-glutamil transpeptidase activity in the kidney of rats

The effect of feeding 24 to 52-days-old male rats of the Wistar strain, with beans diet deficient in sulfur-containing amino acids or supplemented with 0.3% DL-methionine and two corn oil concentrations, 2% and 20% (g/kg/diet), on the glutathione content (GSH) in liver, and on the renal gamma glutamyl transpeptidase (GGTP) (E.C.2.3.2.2) activity was studied. Results indicated that rats fed the bean diet with 2% corn oil decreased significantly the liver GSH content and GGTP renal activity. Supplementation with 0.3% methionine and/or the 20% increment of the diet corn oil, significantly stimulated the liver GSH content and the transpeptidase activity in the kidney. It is postulated that the increase of polyunsaturated fatty acids, stimulated the GGTP activity as a way of increasing substrate bioavailability for synthesis de novo of liver GSH, necessary for the protection of the hydroperoxides formation, attributed to the increment of polyunsaturated acids at cellular level.     

Effect of unusually elevated SO2 atmospheric content on the corrosion of high power electrical conductors - Part 3. Pure copper

This research work belongs to a series of studies simultaneously performed with two previous publications in this journal as Part I. Al and 6201 Al alloy and Part 2. Pure copper. The aim of the project was to have a comparative picture of the joint effect of marine and industrial atmospheric pollutants on the corrosion resistance of wire metals employed for high power electric transmission. This one is also based on the pure Cu behaviour, but limited to six marine-industrial atmospheres with extremely high SO₂ contents. The interest in this study was triggered by unusual results considered appropriate to investigate. Weight loss after 4 and 11 months exposure was determined and morphology of the attack was analysed through SEM-ESEM-EDX. Polarisation of samples after exposure to all the test sites as compared to bare Cu clarified the effect of unusually high SO₂ pollutant contents in these atmospheres on the high protectiveness of the corrosion products formed.     

Evaluation of the adjusted amino acid score by digestibility for estimating the protein quality and protein available in food and diets

The purpose of the present study was to evaluate the amino acid score adjusted by digestibility to estimate protein quality and utilizable protein in foods and diets, considering net protein utilization (NPU) as a biological reference method. Ten foods of vegetable origin and ten of animal origin, as well as eight mixtures of foods of vegetable and animal origin were studied. When all the foods were considered, a positive (r = 0.83) and highly significant correlation (p less than 0.001) between NPU and the amino acid score adjusted by digestibility was found. When the foods were separated according to their origin, this correlation was positive only for the foods of vegetable origin (r = 0.93) and statistically significant (p less than 0.001). Also, only in those foods were similar values found between NPU and amino acid score adjusted by digestibility, as well as in utilizable protein estimated considering both methods. Caution is required to interpret protein quality and utilizable protein values of foods of animal origin and mixtures of foods of vegetable and animal origin when the amino acid score method adjusted by digestibility, or NPU, are utilized.     

Seed-mediated electrochemical growth of gold nanostructures on indium tin oxide thin films

Two-dimensional gold nanostructures (Au NSs) were fabricated on amine-terminated indium tin oxide (ITO) thin films using constant potential electrolysis. By controlling the deposition time and by choosing the appropriate ITO surface, Au NSs with different shapes were generated. When Au NSs were formed directly on aminosilane-modified ITO, the surface roughness of the interface was largely enhanced. Modification of such Au NSs with n-tetradecanethiol resulted in a highly hydrophobic interface with a water contact angle of 144°. Aminosilane-modified ITO films further modified with colloidal Au seeds before electrochemical Au NSs formation demonstrated interesting optical properties. Depending on the deposition time, surface colors ranging from pale pink to beatgold-like were observed. The optical properties and the chemical stability of the interfaces were characterized using UV-vis absorption spectroscopy. Well-defined localized surface plasmon resonance signals were recorded on Au-seeded interfaces with λ_max = 675 ± 2 nm (deposition time 180 s). The prepared interfaces exhibited long-term stability in various solvents and responded linearly to changes in the corresponding refractive indices.     

Hydrolysis of pumpkin oil cake protein isolate and free radical scavenging activity of hydrolysates: Influence of temperature,enzyme/substrate ratio and time

The effect of hydrolysis parameters (temperature, initial enzyme/substrate ratio and time) on the hydrolysis of pumpkin oil cake protein isolate (PuOC PI) with acid protease from Aspergillus niger and the antioxidant potency of the obtained hydrolysates were studied by response surface methodology (RSM). The hydrolysis progress, measured by the degree of hydrolysis (DH), was described by a second-order polynomial model (R² = 0.77) and the conditions for optimum DH (42.94%) were found at temperature of 40 °C, enzyme/substrate ratio (E/S) 4.38 HUT/mg of substrate proteins and 85 min. The antiradical activity (AA) of the PuOC PI hydrolysates was examined by the 1,1-diphenyl-2-picryl-hydrazyl (DPPH) assay; all hydrolysates showed a concentration dependent scavenging activity against DPPH radicals. The AA of hydrolysates was influenced by process parameters and was presented also by a second-order polynomial model (R² = 0.7). The conditions to achieve the highest DH did not result hydrolysates with the optimum AA; the highest AA ranged from 34% to 40% and were found in hydrolysates obtained at 50 °C.