Mapping phosphorylation sites: a new strategy based on the use of isotopically labelled DTT and mass spectrometry

Phosphoproteomics, nowadays, represents a front line in functional proteomics as testified by the number of papers recently appearing in the literature. In an attempt to improve and simplify the methods so far suggested we have set up a simple isotope-coded approach to label and quantitate phospho-Ser/-Thr residues in protein mixtures. First of all, after appropriate oxidation of cysteine/cystine residues followed by tryptic hydrolysis, we have optimised and simplified the beta-elimination reaction to get the corresponding alkene moiety from the phosphate esters. This was achieved by (a) separating the elimination reaction from the addition reaction, (b) the use of Ba(OH)(2) as alkali reagent and (c) its further elimination by the simple addition of solid CO(2) to the peptide mixture. The Michael reaction was then performed, after the removal of BaCO(3) by centrifugation, by adding dithiothreitol (DTT) to the peptide mixture. Finally, the direct purification of the modified phosphopeptides was performed on a thiol-sepharose column. The availability of fully deuterated DTT, introducing a 6 Da difference with respect to the non-deuterated species, allows quantitation of the differential extent of signalling modification when analysed by matrix-assisted laser desorption/ionisation mass spectrometry (MALDI-MS) and liquid chromatography/mass spectrometry. The entire procedure has been set up by using bovine alpha-casein, and resulted in the identification of all the phosphorylated tryptic peptides, including the tetraphosphorylated peptides, which escaped all previously reported procedures     

Measurement of trihalomethanes and methyl tertiary-butyl ether in tap water using solid-phase microextraction GC-MS

The prevalence of water disinfection byproducts in drinking water supplies has raised concerns about possible health effects from chronic exposure to these compounds. To support studies exploring the relation between exposure to trihalomethanes (THMs) and health effects, we have developed an automated analytical method using headspace solid-phase microextraction coupled with capillary gas chromatography and mass spectrometry. This method quantitates trace levels of THMs (chloroform, bromodichloromethane, dibromochloromethane, and bromoform) and methyl tertiary-butyl ether in tap water. Detection limits of less than 100 ng/L for all analytes and linear ranges of three orders of magnitude are adequate for measuring the THMs in tap water samples tested from across the United States. THMs are stable for extended periods in tap water samples after quenching of residual chlorine and buffering to pH 6.5, thus enabling larger epidemiologic field studies with simplified sample collection protocols.     

Truncation of the correlation consistent basis sets: an effective approach to the reduction of computational cost?

The systematic reduction of commonly used basis sets as a means to reduce computational cost is examined for a small test set of molecules, which includes H(2), CH(4), NH(3), H(2)O, HF, and HCN. Coupled cluster with single, double, and quasiperturbative triple excitations calculations were performed using both the correlation consistent basis sets, and a set of systematically reduced basis sets to examine both the impact of the reduction upon the accuracy of the structures and energies, and the computational cost savings achieved. The effect of several truncation scenarios upon basis set convergence is also examined. Overall, for the systems studied, a reduction can occur which preserves the well-established systematic convergence behavior of the correlation consistent basis sets.     

The relation of lung volume initiation to selected acoustic properties of speech

This study examined the relationship of speech breathing to other elements of speech production. It was hypothesized that initiating speech from different lung volumes would have an effect on different elements of the acoustic output. It was postulated that effects may be brought about by mechanical interaction as well as a dispersion of effort to mechanically unlinked elements of speech production, such as articulatory behavior. To this end, selected acoustic variables were studied in eight young healthy women who initiated speech from low, typical, and high lung volume levels. The acoustic variables studied were selected because they have been shown to be sensitive indicators of speech production performance. It was found that with increasing lung volume initiation levels, average sound pressure level, average fundamental frequency, and declination rate of fundamental frequency increased. It was also observed that vowel space was significantly smaller during low lung volume initiation levels relative to typical lung volume initiation levels. Vowel space reduction is discussed relative to "gaining down."     

Wavelet stabilization and preconditioning for domain decomposition

The aim of this paper is to propose a stabilization technique in order to circumvent the two inf–sup conditions neededfor stability of the discretization of the three fields formulationfor domain decomposition methods (following the work of Brezzi& Marini (1993) Optimisation et Controle 89–96). Realization of this technique in terms of wavelets is presented. Furthermore, the resulting discrete problem is shown to admitan asymptotically optimal preconditioner also based on wavelets. Received 24 February, 1999. Revised 7 October, 1999. ^* aivlis@ian.pv.cnr.it; http://www.ian.pv.cnr.it/~aivlis kunoth@iam.un-bonn.de; http://www.iam.uni-bonn.de/~kunoth     

SR XRF and micro-PIXE studies on ancient metallurgy of thirteen Dacian gold bracelets

A thorough microscopic investigation by SR XRF and micro-PIXE brings insight into the probable techniques used in the manufacture of thirteen Dacian gold bracelets, one of the most spectacular archaeological finds ever on the territory of Romania.     

Regular partitions of dual polar spaces

We describe several classes of regular partitions of dual polar spaces and determine their associated parameters and eigenvalues. We also give some general methods for computing the eigenvalues of regular partitions of dual polar spaces.     

Existence and multiplicity results to Neumann problems for elliptic equations involving the p-Laplacian

In this paper, existence results of positive solutions to a Neumann problem involving the p-Laplacian are established. Multiplicity results are also pointed out. The approach is based on variational methods.