Sensitivity-optimized experiment for the measurement of residual dipolar couplings between amide protons

High signal to noise is a necessity for the quantification of NMR spectral parameters to be translated into accurate and precise restraints on protein structure and dynamics. An important source of long-range structural information is obtained from ¹H–¹H residual dipolar couplings (RDCs) measured for weakly aligned molecules. For sensitivity reasons, such measurements are generally performed on highly deuterated protein samples. Here we show that high sensitivity is also obtained for protonated protein samples if the pulse schemes are optimized in terms of longitudinal relaxation efficiency and J-mismatch compensated coherence transfer. The new sensitivity-optimized quantitative J-correlation experiment yields important signal gains reaching factors of 1.5 to 8 for individual correlation peaks when compared to previously proposed pulse schemes. Paul Schanda and Ewen Lescop contributed equally to this work.     

地下水生态补偿标准核算模型

地下水是水资源的重要组成部分,在维持生产生活、推动社会经济发展方面具有重要作用。地下水生态补偿有助于促进地下水资源合理开发利用,但是国内外关于此方面尤其是补偿标准的研究非常缺乏。本研究提出由基础、奖惩、科技和风险补偿构成的地下水生态补偿标准核算模型,综合考虑气候条件和经济水平,明确补偿优先等级,并将模型应用于山西省11个市,核算其地下水生态补偿标准。结果表明: 基础补偿在总补偿中占比最大,且非市场价值占比在60%以上,说明地下水具有较高的调节服务价值;2008—2017年,各市发展系数有明显提高,说明区域经济水平提升,补偿能力有所增强;补偿优先等级受地下水非市场价值及区域经济水平的影响,各地补偿优先等级差异明显,需要尽快落实各地的地下水生态补偿工作。同时,建议尽快完善地下水风险补偿制度,设立地下水生态科技补偿专项资金,建立长期有效的地下水生态补偿机制。     

The effects of charge compensation on photoluminescence properties of a new green‐emitting ZnB2O4:Tb3+ phosphor

Charge compensation is an effective way to eliminate charge defects and improve the luminescent intensity of phosphors. In this paper, a new green‐emitting phosphor ZnB₂O₄:Tb³⁺ was prepared by solid‐state reaction at 750°C. The effects of Tb³⁺ doping content and charge compensators (Li⁺, Na⁺ or K⁺) on photoluminescence properties of ZnB₂O₄:Tb³⁺ were investigated. X‐ray powder diffraction analysis confirms the sample has cubic structure of ZnB₂O₄. The excitation and emission spectra indicate that this phosphor can be excited by near ultraviolet light at 378 nm, and exhibits bright green emission with the highest peak at 544 nm corresponding to the ⁵D₄→⁷F₅ transition of Tb³⁺. The critical quenching concentration of Tb³⁺ in ZnB₂O₄ host is 8 mol%. The results of charge compensation show that the emission intensity can be improved by Na⁺ and K⁺. Specifically, K⁺ is the optimal one for ZnB₂O₄:Tb³⁺. Copyright © 2014 John Wiley & Sons, Ltd.     

A targeted proteomics toolkit for high-throughput absolute quantification of Escherichia coli proteins

Transformation of engineered Escherichia coli into a robust microbial factory is contingent on precise control of metabolism. Yet, the throughput of omics technologies used to characterize cell components has lagged far behind our ability to engineer novel strains. To expand the utility of quantitative proteomics for metabolic engineering, we validated and optimized targeted proteomics methods for over 400 proteins from more than 20 major pathways in E. coli metabolism. Complementing these methods, we constructed a series of synthetic genes to produce concatenated peptides (QconCAT) for absolute quantification of the proteins and made them available through the Addgene plasmid repository (www.addgene.org). To facilitate high sample throughput, we developed a fast, analytical-flow chromatography method using a 5.5-min gradient (10 min total run time). Overall this toolkit provides an invaluable resource for metabolic engineering by increasing sample throughput, minimizing development time and providing peptide standards for absolute quantification of E. coli proteins.     

Anthropometric and fitness normative values for young karatekas

To provide information regarding the anthropometric and fitness profile of young karatekas and to study its evolution with age. Data from top-level karatekas were included in the analysis: 97 athletes in the U14 category (12–13 years old), 238 in cadet (14–15 years old), 261 in junior (16–17 years old) and 177 in U21 (18–20 years old), which makes a total of 773 athlete data sets. Karatekas underwent anthropometric (weight, height, body mass index and body fat percentage) and fitness (sit and reach, 20-m shuttle run, standing long jump, overhead 3-kg ball throw, 10x5-m shuttle run, and plate-tapping) assessments during the training camps organized by the Spanish National Karate Federation between 1999 and 2016. Male karatekas were taller and heavier, and performed better than females in all the fitness dimensions assessed, except for flexibility and speed of upper limb movements. The obtained cardiovascular and lower-body muscular values indicated that karatekas in this study were placed between the 80th and the 90th percentile when compared with the general population. The results of the agility, coordination and speed of upper limb movements, and flexibility tests showed that the karatekas obtained much higher scores than those observed in age-matched populations. Young karatekas show a high fitness level in comparison with the general population, especially with regards to aerobic performance, lower-body muscular power and upper-limb movement speed. Reference values of anthropometric and fitness dimensions are provided in order to be used by coaches, conditioning trainers and sport scientists when testing young male and female karatekas.     

Methylation and expression analyses of Pallister-Killian syndrome reveal partial dosage compensation of tetrasomy 12p and hypomethylation of gene-poor regions on 12p

To ascertain the epigenomic features, i.e., the methylation, non-coding RNA, and gene expression patterns, associated with gain of i(12p) in Pallister-Killian syndrome (PKS), we investigated single cell clones, harboring either disomy 12 or tetrasomy 12p, from a patient with PKS. The i(12p)-positive cells displayed a characteristic expression and methylation signature. Of all the genes on 12p, 13% were overexpressed, including the ATN1, COPS7A, and NECAP1 genes in 12p13.31, a region previously implicated in PKS. However, the median expression fold change (1.3) on 12p was lower than expected by tetrasomy 12p. Thus, partial dosage compensation occurs in cells with i(12p). The majority (89%) of the significantly deregulated genes were not situated on 12p, indicating that global perturbation of gene expression is a key pathogenetic event in PKS. Three genes—ATP6V1G1 in 9q32, GMPS in 3q25.31, and TBX5 in 12q24.21—exhibited concomitant hypermethylation and decreased expression. The i(12p)-positive cells displayed global hypomethylation of gene-poor regions on 12p, a footprint previously associated with constitutional and acquired gains of whole chromosomes as well as with X-chromosome inactivation in females. We hypothesize that this non-genic hypomethylation is associated with chromatin processing that facilitates cellular adaptation to excess genetic material.     

Quantitative analysis of sphingolipids for lipidomics using triple quadrupole and quadrupole linear ion trap mass spectrometers

Sphingolipids are a highly diverse category of bioactive compounds. This article describes methods that have been validated for the extraction, liquid chromatographic (LC) separation, identification and quantitation of sphingolipids by electrospray ionization, tandem mass spectrometry (ESI-MS/MS) using triple quadrupole (QQQ, API 3000) and quadrupole-linear-ion trap (API 4000 QTrap, operating in QQQ mode) mass spectrometers. Advantages of the QTrap included: greater sensitivity, similar ionization efficiencies for sphingolipids with ceramide versus dihydroceramide backbones, and the ability to identify the ceramide backbone of sphingomyelins using a pseudo-MS³ protocol. Compounds that can be readily quantified using an internal standard cocktail developed by the LIPID MAPS Consortium are: sphingoid bases and sphingoid base 1-phosphates, more complex species such as ceramides, ceramide 1-phosphates, sphingomyelins, mono- and di-hexosylceramides, and these complex sphingolipids with dihydroceramide backbones. With minor modifications, glucosylceramides and galactosylceramides can be distinguished, and more complex species such as sulfatides can also be quantified, when the internal standards are available. JLR LC ESI-MS/MS can be utilized to quantify a large number of structural and signaling sphingolipids using commercially available internal standards. The application of these methods is illustrated with RAW264.7 cells, a mouse macrophage cell line. These methods should be useful for a wide range of focused (sphingo)lipidomic investigations.     

Ecological opportunity and the origin of adaptive radiations

Ecological opportunity – through entry into a new environment, the origin of a key innovation or extinction of antagonists – is widely thought to link ecological population dynamics to evolutionary diversification. The population‐level processes arising from ecological opportunity are well documented under the concept of ecological release. However, there is little consensus as to how these processes promote phenotypic diversification, rapid speciation and adaptive radiation. We propose that ecological opportunity could promote adaptive radiation by generating specific changes to the selective regimes acting on natural populations, both by relaxing effective stabilizing selection and by creating conditions that ultimately generate diversifying selection. We assess theoretical and empirical evidence for these effects of ecological opportunity and review emerging phylogenetic approaches that attempt to detect the signature of ecological opportunity across geological time. Finally, we evaluate the evidence for the evolutionary effects of ecological opportunity in the diversification of Caribbean Anolis lizards. Some of the processes that could link ecological opportunity to adaptive radiation are well documented, but others remain unsupported. We suggest that more study is required to characterize the form of natural selection acting on natural populations and to better describe the relationship between ecological opportunity and speciation rates.     

完善公立医院补偿机制改革的难点与思考

补偿机制改革是公立医院改革的重点和难点。在分析当前公立医院财政补助现状以及取消药品加成的影响的基础上,指出当前补偿途径在具体实施过程中还存在相当的困难。为保障公立医院补偿机制改革的顺利推进,应从医疗服务的公平性和效益出发,明确补偿范围,制定补偿标准,并在机制体制上确保补偿到位。