Backbone and sidechain 1H, 13C and 15N resonance assignments of the RGS domain from human RGS14

We have assigned ¹H, ¹⁵N and ¹³C resonances of the RGS domain from the human RGS14 protein, a multi-domain member of the RGS (Regulators of G-protein signalling) family of proteins, important in the down-regulation of specific G-protein signalling pathways.     

An ant colony optimization approach to flexible protein–ligand docking

The prediction of the complex structure of a small ligand with a protein, the so-called protein–ligand docking problem, is a central part of the rational drug design process. For this purpose, we introduce the docking algorithm PLANTS (Protein–Ligand ANT System), which is based on ant colony optimization, one of the most successful swarm intelligence techniques. We study the effectiveness of PLANTS for several parameter settings and present a direct comparison of PLANTS’s performance to a state-of-the-art program called GOLD, which is based on a genetic algorithm and frequently used in the pharmaceutical industry for this task. Last but not least, we also show that PLANTS can make effective use of protein flexibility giving example results on cross-docking and virtual screening experiments for protein kinase A. This article is based on a paper that won the best paper award at ANTS 2006, the 5th International Workshop on Ant Colony Optimization and Swarm Intelligence held in Brussels, Belgium, 2006. This article includes new types of experiments and also the possibility of considering flexibility of protein side-chains.     

Blood pressure and hypertension in an American colony (Puerto Rico) and on the USA mainland compared, 1886–1930

We compare blood pressure and hypertension between adult men on the USA mainland and in Puerto Rico born during 1886-1930 to test hypotheses about the link between cardiovascular health and large socioeconomic and political changes in society: (a) 8853 men surveyed in Puerto Rico in 1965 and (b) 1449 non-Hispanic White men surveyed on the mainland during 1971-1975. Systolic and diastolic blood pressure and hypertension were regressed separately on demographic and socioeconomic variables and cardiovascular risk factors. Mainland men not taking anti-hypertensive medication showed statistically significant improvements in systolic blood pressure and hypertension at the beginning of the century and men in Puerto Rico showed improvements in diastolic blood pressure but only during the last two quinquenniums. An average man born on the mainland during the last birth quinquennium (1926-1930) had 7.4-8.7 mmHg lower systolic blood pressure and was 61% less likely to have systolic hypertension than one born before 1901. On average Puerto Rican men born during 1921-1925 had approximately 1.7 mmHg lower diastolic blood pressure than men born before 1901. Analyses of secular trends in cardiovascular health complements analyses of secular trends in anthropometric indicators and together provide a fuller view of the changing health status of a population.     

Simultaneous quantification of voriconazole and posaconazole in human plasma by high-performance liquid chromatography with ultra-violet detection

A sensitive and selective high-performance liquid chromatographic (HPLC) method with ultra-violet detection has been developed and validated for the simultaneous determination of posaconazole and voriconazole, two systemic anti-fungal agents. An internal standard diazepam was added to 100 microL of human plasma followed by 3 mL of hexane-methylene chloride (70:30, v/v). The organic layer was evaporated to dryness and the residue was reconstituted with 100 microL of mobile phase before being injected in the chromatographic system. The compounds were separated on a C8 column using sodium potassium phosphate buffer (0.04 M, pH 6.0): acetonitrile:ultrapure water (45:52.5:2.5, v/v/v) as mobile phase. All compounds were detected at a wavelength of 255 nm. The assay was linear and validated over the range 0.2-10.0 mg/L for voriconazole and 0.05-10.0 mg/L for posaconazole. The biases were comprised between -3 and 5% for voriconazole and -2 and 8% for posaconazole. The intra- and inter-day precisions of the method were lower than 8% for the routine quality control (QC). The mean recovery was 98% for voriconazole and 108% for posaconazole. This method provides a useful tool for therapeutic drug monitoring.     

The determination of organophosphonate nerve agent metabolites in human urine by hydrophilic interaction liquid chromatography tandem mass spectrometry

A sensitive, robust isotope dilution LC/MS/MS method is presented for the quantitative analysis of human urine for the alkyl methylphosphonic acid metabolites of five organophosphorus nerve agents (VX, rVX or VR, GB or Sarin, GD or Soman, and GF or Cyclosarin). The selective sample preparation method employs non-bonded silica solid-phase extraction and is partially automated. While working with a mobile phase composition that enhances the electrospray ionization process, the hydrophilic interaction chromatography method results in a 5-min injection-to-injection cycle time, excellent peak shapes and adequate retention (k'=3.1). These factors lead to limits of detection for these metabolites as low as 30 pg/mL in a 1-mL sample of human urine. The quality control data (15 and 75 ng/mL) demonstrate accurate (-0.5 to +3.4%) and precise (coefficients of variation of 2.1-3.6%) quantitative results over the clinically relevant urine concentration range of 1-200 ng/mL for a validation set of 20 standard and quality control sets prepared by five analysts over 54 days. The selectivity of the method is demonstrated for a 100-individual reference range study, as well as the analysis of relevant biological samples. The combined sample preparation and analysis portions of this method have a throughput of 288 samples per day.     

A rapid and sensitive method for the quantitation of montelukast in sheep plasma using liquid chromatography/tandem mass spectrometry

A rapid LC-MS/MS method was developed and partially validated for the quantitation of montelukast in spiked sheep plasma. A total run time of 1.5 min was achieved using a short monolithic column and employing a rapid gradient. Sample preparation involved protein precipitation with twofold acetonitrile by volume during which a deuterated internal standard (montelukast D-6) was incorporated. The MRM transitions for montelukast and the deuterated internal standard were 586/422 and 592/427, respectively. A linear dynamic range of 0.25-500 ng/mL with a correlation coefficient of 0.9999 was achieved. Precision was below 5% at all levels except at the LOQ (0.36 ng/mL) which demonstrated an overall of R.S.D. of 8%. Post-column infusion experiments were performed with precipitated plasma matrix and showed minimal interference with the peaks of interest.     

Impact of Land-use Change on Dengue and Malaria in Northern Thailand

Land-use change, a major constituent of global environmental change, potentially has significant consequences for human health in relation to mosquito-borne diseases. Land-use change can influence mosquito habitat, and therefore the distribution and abundance of vectors, and land use mediates human–mosquito interactions, including biting rate. Based on a conceptual model linking the landscape, people, and mosquitoes, this interdisciplinary study focused on the impacts of changes in land use on dengue and malaria vectors and dengue transmission in northern Thailand. Extensive data on mosquito presence and abundance, land-use change, and infection risk determinants were collected over 3 years. The results of the different components of the study were then integrated through a set of equations linking land use to disease via mosquito abundance. The impacts of a number of plausible scenarios for future land-use changes in the region, and of concomitant behavioral change were assessed. Results indicated that land-use changes have a detectable impact on mosquito populations and on infection. This impact varies according to the local environment but can be counteracted by adoption of preventive measures.     

SPINE: a method for the rapid detection and analysis of protein-protein interactions in vivo

The detection and analysis of protein-protein interactions is one of the central tasks of proteomics in the postgenomic era. For this purpose, we present a procedure, the Strep-protein interaction experiment (SPINE) that combines the advantages of the Strep-tag protein purification system with those of reversible in vivo protein crosslinking by formaldehyde. Using two Bacillus subtilis regulator proteins, we demonstrate that this method is well suited to isolate protein complexes with high purity and virtually no background. Plasmids allowing the high-level expression of proteins carrying an N- or C-terminal Strep-tag in B. subtilis were constructed.     

Effects of ionizing radiation on the activity of the major hepatic enzymes implicated in bile acid biosynthesis in the rat

In the days following high-dose radiation exposure, damage to small intestinal mucosa is aggravated by changes in the bile acid pool reaching the gut. Intestinal bile acid malabsorption, as described classically, may be associated with altered hepatic bile acid biosynthesis, which was the objective of this work. The activity of the main rate-limiting enzymes implicated in the bile acid biosynthesis were evaluated in the days following an 8-Gy gamma(60)Co total body irradiation of rats, with concomitant determination of biliary bile acid profiles and intestinal bile acid content. Modifications of biliary bile acid profiles, observed as early as the first post-irradiation day, were most marked at the third and fourth day, and resulted in an increased hydrophobicity index. In parallel, the intestinal bile acids' content was enhanced and hepatic enzymatic activities leading to bile acids were changed. A marked increase of sterol 12 alpha-hydroxylase and decrease of oxysterol 7 alpha-hydroxylase activity was observed at day 3, whereas both cholesterol 7 alpha-hydroxylase and oxysterol 7 alpha-hydroxylase activities were decreased at day 4 after irradiation. These results show, for the first time, radiation-induced modifications of hepatic enzymatic activities implicated in bile acid biosynthesis and suggest that they are mainly a consequence of radiation-altered intestinal absorption, which induces a physiological response of the enterohepatic bile acid recirculation.     

Malaria Plasmodium agent induces alteration in the head proteome of their Anopheles mosquito host

Despite increasing evidence of behavioural manipulation of their vectors by pathogens, the underlying mechanisms causing infected vectors to act in ways that benefit pathogen transmission remain enigmatic in most cases. Here, 2-D DIGE coupled with MS were employed to analyse and compare the head proteome of mosquitoes (Anopheles gambiae sensu stricto (Giles)) infected with the malarial parasite (Plasmodium berghei) with that of uninfected mosquitoes. This approach detected altered levels of 12 protein spots in the head of mosquitoes infected with sporozoites. These proteins were subsequently identified using MS and functionally classified as belonging to metabolic, synaptic, molecular chaperone, signalling, and cytoskeletal groups. Our results indicate an altered energy metabolism in the head of sporozoite-infected mosquitoes. Some of the up-/down-regulated proteins identified, such as synapse-associated protein, 14-3-3 protein and calmodulin, have previously been shown to play critical roles in the CNS of both invertebrates and vertebrates. Furthermore, a heat shock response (HSP 20) and a variation of cytoarchitecture (tropomyosins) have been shown. Discovery of these proteins sheds light on potential molecular mechanisms that underlie behavioural modifications and offers new insights into the study of intimate interactions between Plasmodium and its Anopheles vector.