Copper(II) interaction with unstructured prion domain outside the octarepeat region: speciation, stability, and binding details of copper(II) complexes with PrP106-126 peptides

Copper(II) complexes of the neurotoxic peptide fragments of human and chicken prion proteins were studied by potentiometric, UV-vis, CD, and EPR spectroscopic and ESI-MS methods. The peptides included the terminally blocked native and scrambled sequences of HuPrP106-126 (HuPrPAc106-126NH2 and ScrHuPrPAc106-126NH2) and also the nona- and tetrapeptide fragments of both the human and chicken prion proteins (HuPrPAc106-114NH2, ChPrPAc119-127NH2, HuPrPAc109-112NH2, and ChPrPAc122-125NH2). The histidyl imidazole-N donor atoms were found to be the major copper(II) binding sites of all peptides; 3N and 4N complexes containing additional 2 and 3 deprotonated amide-N donors, respectively, are the major species in the physiological pH range. The complex formation processes for nona- and tetrapeptides are very similar, supporting the fact that successive deprotonation and metal ion coordination of amide functions go toward the N-termini in the form of joined six- and five-membered chelates. As a consequence, the peptide sequences investigated here, related to the neurotoxic region of the human PrP106-126 sequence, show a higher metal-binding affinity than the octarepeat fragments. In the case of the HuPrP peptide sequences, a weak pH-dependent binding of the Met109 residue was also detected in the 3N-coordinated complexes.     

A proteomic approach to study protein variation in GM durum wheat in relation to technological properties of semolina

Genetic manipulation of durum wheats by tobacco rab-1 genes influence the trafficking of gluten proteins through the secretory system by up- or down-regulating the transport step from the ER to the Golgi apparatus which may in turn modify functional performance of the grain. Gluten proteins were extracted from two genetically manipulated lines - Svevo B730 1-1 and Ofanto B688 1-2 - and their control lines and were analyzed by two dimensional gel electrophoresis. When the two-dimensional maps were compared by image analysis no significant differences between the GM line with an up-regulated trafficking containing the wild type tobacco rab1 (Svevo B730 1-1) and its control (Svevo control). By contrast, significant differences were found between the GM line with a down-regulated trafficking due to the tobacco rab1 mutant form (Ofanto B688 1-2) and its control (Ofanto control). Of the new protein spots detected in the down-regulated Ofanto B688 1-2 map, only a beta-amylase was identified. The remaining spots were susceptible to chymotripsin action but not to trypsin one, as in the case of the gluten protein. Rheological measurements showed that gluten quality was enhanced in the down-regulated Ofanto B688 1-2 without an increase in the amount of gluten. Proteomics is a useful and powerful tool for investigating protein changes in GMOs and in understanding events in food science and technology.     

Simultaneous determination of phenytoin and dextromethorphan in urine by solid-phase extraction and HPLC-DAD

A rapid and simple high-performance liquid chromatographic method with photodiode array detection was developed for the separation and the simultaneous determination of phenytoin and dextromethorphan in human urine. Analysis was performed in less than 4.5 min in isocratic mode on a reversed-phase C18 column (5 microm; 150 x 4.6 mm) using a mobile phase composed of acetonitrile-buffer phosphate 0.01 M (60:40, v/v) adjusted to pH 6.0, at 1 mL/min flow rate and UV absorbance at 210 nm. The elution order of analytes was dextromethorphan (DXM), Internal Standard (IS), and phenytoin (PHT). Calibration curves were linear in the 7.5-25 microg/mL range for PHT and in the 10-30 microg/mL range for DXM. Spike recoveries for urine samples prepared at three spiking levels ranged from 97.8 to 102.3% for PHT and from 94.8 to 100.4% for DXM. The detection limit (LOD) values ranged from 0.08 microg/mL for PHT to 0.5 microg/mL for DXM. The quantitation limit (LOQ) values ranged from 0.3 microg/mL for PHT to 1.6 microg/mL for DXM. The sample preparation method involves a rapid and simple procedure based on solid-phase extraction using a C18 reversed-phase column. Validation of the optimised method was carried out according to the ICH guidelines. The method developed in this study allows the reliable simultaneous analysis of PHT and DXM, drugs that were never quantified together in previously reported analytical methods. The described method has the advantage of being rapid and easy and it could be applied in therapeutic monitoring of these drugs in human urine of epileptic patients.     

Wave-Like Solutions for a Class of Parabolic Models

A reduction aproach is developed for determining exact solutions of anonlinear second order parabolic PDE. The method in point makes acomplementary use of the leading ideas of the theory of quasilinearhyperbolic systems of first order endowed by differential constraintsand of the techniques providing multiple wave-like solutions ofnonlinear PDEs. The searched solutions exhibit a inherent wave featuresand they are obtained by solving a consistent overdetermined system ofPDEs. Remarkably, in the process it is possible to define nonlinearmodel equations which allow special classes of initial or boundary valueproblems to be solved in a closed form. Within the present reductionapproach exact solutions and model material response functions areobtained for an equation of widespread application in many fields ofinterest.