Determination of phosphorus-, copper-, and zinc-containing human brain proteins by LA-ICPMS and MALDI-FTICR-MS

Human brain proteins containing phosphorus, copper, and zinc were detected directly in protein spots in gels of a human brain sample after separation by two-dimensional gel electrophoresis using laser ablation inductively coupled plasma mass spectrometry (LA-ICPMS). A powerful laser ablation system with cooled laser ablation chamber was coupled to a double-focusing sector field ICPMS. The separated protein spots in 2D gels were fast screened using the optimized microanalytical LA-ICPMS technique measured at medium mass resolution with a focused laser beam (wavelength, 213 nm; diameter of laser crater, 50 mum; and laser power density, 3 x 10(9) W cm(-2)) with respect to selected three essential elements. Of 176 protein spots in 2D gel from a human brain sample, phosphorus, copper, and zinc were detected in 31, 43, and 49 protein spots, respectively. For the first time, uranium as a naturally occurring radioactive element was found in 20 selected protein spots. The detection limits for P, S, Cu, Zn and U were determined in singular protein spots with 0.0013, 1.29, 0.029, 0.063, and 0.000 01 mg g(-1), respectively. A combination of LA-ICPMS with matrix-assisted laser desorption/ionization Fourier transform ion cyclotron resonance mass spectrometry (MALDI-FTICR-MS) was applied for the identification of selected protein spots from human brain protein separated by 2D gel electrophoresis. Combining MALDI-FTICR-MS for the structure analysis of metal- and phosphorus-containing human brain proteins with LA-ICPMS, the direct analysis of heteroelements on separated proteins in 2D gels can be performed. For quantification of analytical LA-ICPMS data, the number of sulfur atoms per protein (and following the sulfur concentration) determined by MALDI-FTICR-MS was used for internal standardization. From the known sulfur concentration in protein, the concentration of other heteroelements was calculated. In addition, the number of phosphorylation and the phosphorylation sites of phosphorylated proteins in the human brain sample detected by LA-ICPMS were determined by MALDI-FTICR-MS. This technique allows the study of posttranslational modifications in human brain proteins.     

Radiation transport calculations for 50 MV photon therapy beam using the Monte Carlo code GEANT4

A new thin transmission target technique for fast dose delivery using narrow scanned photon beams has been developed. High-energy, 50-100 MeV, electron beams of low emittance incident on thin low-Z targets produce narrow and intense high-energy bremsstrahlung beams. However, electrons transmitted through the target are bent from the therapeutic beam by a purging magnet and have to be effectively absorbed in a dedicated electron collector. The electron-photon transport through a treatment head has been studied using the Monte Carlo simulation toolkit Geant4. The Geant4 electromagnetic physics processes have been compared with experimental data of radial dose profiles. The differences between calculated and measured radial dose distributions are approximately 2-10%. Preliminary investigations of the collector design have been carried out in order to minimise secondary electron and photon contamination of the therapeutic beam. The toolkit presented here is promising for further development of narrow photon beam therapy.     

Multivalent HSA conjugates of 3'-sialyllactose are potent inhibitors of adenoviral cell attachment and infection

Adenoviruses of serotypes 8, 19 and 37 are the major cause of the severe eye infection EKC (epidemic keratoconjunctivitis). In general, all adenoviruses interact with their cellular receptors through the fibre proteins, which extend from the virus particle. Recently, adenovirus type 37 (Ad37) was found to bind and infect human corneal cells through attachment to carbohydrate structures that carry terminal alpha-(2-3)-linked sialic acids. Herein we present a synthetic route to a 3'-sialyllactose derivative and corresponding multivalent HSA conjugates with varying orders of valency. The potential of these compounds as inhibitors of EKC-causing adenovirus of serotype Ad37, was studied with both a binding assay and an infectivity assay. The results revealed that these compounds effectively prevent Ad37 from binding to and infecting human corneal epithelial (HCE) cells. Moreover, the inhibition is significantly increased with higher orders of multivalency.     

Beitr?ge zur Biochemie der K?sereifung

Zusammenfassung In Reifungsansätzen von Sauermilchkäse (Harzer Käse) wurde für Proben, die mit normalen Richtsalzzusätzen, mit übergroßen Salzgaben und ohne jeglichen Salzzusatz hergestellt waren, die Argininbilanz verschiedener Stickstofffraktionen der Gesamtkäse-Stickstoffsubstanz ermittelt. Es ergab sich, daß bei normaler Käsebereitung geringe Argininverluste auftreten, daß außerdem freies Arginin nicht oder nur spurenweise im Käse zu finden ist. Daraus wird gefolgert, daß Sauermilch-Käse normaler Herstellung keine Minderung der biologischen Wertigkeit hinsichtlich des Arginins erfahren und Arginin nicht an der Geschmacksausbildung solcher Käse beteiligt ist. Auch für Käse, der im Schnellreifungsverfahren gewonnen wurde, gelten diese Gesichtspunkte. Reifung von Quark ohne jeden Salzzusatz liefert grundsätzlich andere Ergebnisse; derartige, mehr oder minder genußuntaugliche Produkte sind durch hohen Gehalt an freiem Arginin gekennzeichnet, der an der Ausbildung unerwünschter, widerlicher Geschmacksnuancen mitbeteiligt ist.     

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Ohne Zusammenfassung     

Pseudo-Starvation Driven Energy Expenditure Negatively Affects Ovarian Follicle Development

In the present investigation, we examined whether a change in whole body energy fluxes could affect ovarian follicular development, employing mice ectopically expressing uncoupling protein 1 in skeletal muscle (UCP1-TG). Female UCP1-TG and wild-type (WT) mice were dissected at the age of 12 weeks. Energy intake and expenditure, activity, body weight and length, and body composition were measured. Plasma insulin, glucose, leptin, plasma fibroblast growth factor 21 (FGF21) and plasma insulin-like growth factor 1 (IGF1) levels were analyzed and ovarian follicle and corpus luteum numbers were counted. IGF1 signaling was analyzed by immunohistochemical staining for the activation of insulin receptor substrate 1/2 (IRS1/2) and AKT. UCP1-TG female mice had increased energy expenditure, reduced body size, maintained adiposity, and decreased IGF1 concentrations compared to their WT littermates, while preantral and antral follicle numbers were reduced by 40% and 60%, respectively. Corpora lutea were absent in 40% of the ovaries of UCP1-TG mice. Phospho-IRS1, phospho-AKT -Ser473 and -Thr308 immunostaining was present in the granulosa cells of antral follicles in WT ovaries, but faint to absent in the antral follicles of UCP1-TG mice. In conclusion, the reduction in circulating IGF1 levels due to the ectopic expression of UCP1 is associated with reduced immunostaining of the IRS1-PI3/AKT pathway, which may negatively affect ovarian follicle development and ovulation.     

Chemically and enzymatically catalyzed synthesis of C6–C10 alkyl benzoates

The esterification of benzoic acid with n‐hexanol, n‐octanol, 2‐ethyl hexanol and n‐decanol was investigated in detail. An analysis of the reaction kinetics of esterification in the presence of different commercially available chemical catalysts was carried out. The effects of catalyst type and loading on the reaction rate were studied. Although the considered reaction is bimolecular, it showed a first‐order behavior, and a linear dependence with respect to the catalyst concentration was observed. Hence, a new approach is presented to describe the reaction kinetics accurately over a wide range. The application of biotechnological synthesis applying different enzymes as catalysts offers an interesting alternative besides chemical synthesis. Especially an esterase from Bacillus subtilis immobilized on Sepabeads EC‐EP showed high stability and was applied for 2 days in the synthesis of hexyl benzoate. Nevertheless, the chemical reaction route remains superior with respect to the catalyst activities under the applied conditions, which were 25 kU/g for the chemical reaction and 0.7 kU/g for the best enzymatic conversion.