Trypsiligase-Catalyzed Labeling of Proteins on Living Cells

Fluorescent fusion proteins are powerful tools for studying biological processes in living cells, but universal application is limited due to the voluminous size of those tags, which might have an impact on the folding, localization or even the biological function of the target protein. The designed biocatalyst trypsiligase enables site-directed linkage of small-sized fluorescence dyes on the N terminus of integral target proteins located in the outer membrane of living cells through a stable native peptide bond. The function of the approach was tested by using the examples of covalent derivatization of the transmembrane proteins CD147 as well as the EGF receptor, both presented on human HeLa cells. Specific trypsiligase recognition of the site of linkage was mediated by the dipeptide sequence Arg-His added to the proteins’ native N termini, pointing outside the cell membrane. The labeling procedure takes only about 5 minutes, as demonstrated for couplings of the fluorescence dye tetramethyl rhodamine and the affinity label biotin as well.     

Cell-Free Synthesis of Selenoproteins in High Yield and Purity for Selective Protein Tagging

The selenol group of selenocysteine is much more nucleophilic than the thiol group of cysteine. Selenocysteine residues in proteins thus offer reactive points for rapid post-translational modification. Herein, we show that selenoproteins can be expressed in high yield and purity by cell-free protein synthesis by global substitution of cysteine by selenocysteine. Complete alkylation of solvent-exposed selenocysteine residues was achieved in 10 minutes with 4-chloromethylene dipicolinic acid (4Cl-MDPA) under conditions that left cysteine residues unchanged even after overnight incubation. Gd^III−Gd^III distances measured by double electron–electron resonance (DEER) experiments of maltose binding protein (MBP) containing two selenocysteine residues tagged with 4Cl-MDPA-Gd^III were indistinguishable from Gd^III−Gd^III distances measured of MBP containing cysteine reacted with 4Br-MDPA tags.     

A probabilistic load shedding concept considering highly volatile local generation

Automatic load shedding is the ultimate countermeasure against imbalance in a power system and can effectively help preventing large blackouts. Taking into account a high penetration of renewable energy sources (RES) in the distribution grid, a clear distinction between load and generation at the PCC becomes increasingly more difficult. For that reason an adaptation of frequency relay parameters and their locations of installation are necessary. In Europe this is rest on a multi-step plan based on values such as the yearly peak load. In this paper a novel probabilistic method for automatic load shedding is presented that uses the average values instead of peak values for load shedding. Its applicability is verified by a dynamic power system model that was developed to compare the classical and novel probabilistic load shedding principle. The method is verified using data from a German TSO.     

Selective Analysis of Sulfur-Containing Species in a Heavy Crude Oil by Deuterium Labeling Reactions and Ultrahigh Resolution Mass Spectrometry

A heavy crude oil has been treated with deuterated alkylating reagents (CD₃I and C₂D₅I) and directly analyzed without any prior fractionation and chromatographic separation by high-field Orbitrap Fourier Transform Mass Spectrometry (FTMS) and Fourier Transform Ion Cyclotron Resonance Mass Spectrometry (FT-ICR MS) using electrospray ionization (ESI). The reaction of a polycyclic aromatic sulfur heterocycles (PASHs) dibenzothiophene (DBT), in the presence of silver tetrafluoroborate (AgBF₄) with ethyl iodide (C₂H₅I) in anhydrous dichloroethane (DCE) was optimized as a sample reaction to study heavy crude oil mixtures, and the reaction yield was monitored and determined by proton nuclear magnetic resonance spectroscopy (¹H-NMR). The obtained conditions were then applied to a mixture of standard aromatic CH-, N-, O- and S-containing compounds and then a heavy crude oil, and only sulfur-containing compounds were selectively alkylated. The deuterium labeled alkylating reagents, iodomethane-d₃ (CD₃I) and iodoethane-d₅ (C₂D₅I), were employed to the alkylation of heavy crude oil to selectively differentiate the tagged sulfur species from the original crude oil.     

恒星光干涉仪样机的总控系统

控制系统以SCO公司的UNIX ODT为软件平台,普通PC机为总控计算机,通用8098单 片机系统作为前端机组成一个主从式两级管理系统,与PC机相连的16通道多用户卡通过RS-232与14个前端机通讯,进行实时控制和处理。用C语言编写了该样机的总控软件,前端机的通讯程序和相关控制部件的控制程序。经过调试,实现了对样机的全自动控制。     

RFA—300型自动分析仪检测总植物碱的改进

对连续流动法测定烟草及烟草制品中总植物碱的各影响因素进行了试验研究,并根据试验结果对仪器管路进行相应的改造。实验结果表明,烟草制品中总植物碱的连续流动法测定,改进后方法的回收收率高、重现性好。     

对水轮机自动调速器油压控制电路的改进

YT－1000型油压自动调速器在运行中不能维持正常油压、在不增加任何设备的情况下，只改动二次接线的处理方法，就能消除隐患、解决问题。     

地下商场空调设计问题的探讨

本文分析了地下商场的特征 ,并相应提出了地下商场通风空调设计的有关数据、方法及注意事项