Soluble Prokaryotic Overexpression and Purification of Human GM-CSF Using the Protein Disulfide Isomerase b′a′ Domain

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a member of the colony-stimulating factor (CSF) family, which functions to enhance the proliferation and differentiation of hematopoietic stem cells and other hematopoietic lineages such as neutrophils, dendritic cells, or macrophages. These proteins have thus generated considerable interest in clinical therapy research. A current obstacle to the prokaryotic production of human GM-CSF (hGM-CSF) is its low solubility when overexpressed and subsequent complex refolding processes. In our present study, the solubility of hGM-CSF was examined when combined with three N-terminal fusion tags in five E. coli strains at three different expression temperatures. In the five E. coli strains BL21 (DE3), ClearColi BL21 (DE3), LOBSTR, SHuffle T7 and Origami2 (DE3), the hexahistidine-tagged hGM-CSF showed the best expression but was insoluble in all cases at each examined temperature. Tagging with the maltose-binding protein (MBP) and the b′a′ domain of protein disulfide isomerase (PDIb′a′) greatly improved the soluble overexpression of hGM-CSF at 30 °C and 18 °C. The solubility was not improved using the Origami2 (DE3) and SHuffle T7 strains that have been engineered for disulfide bond formation. Two conventional chromatographic steps were used to purify hGM-CSF from the overexpressed PDIb′a′-hGM-CSF produced in ClearColi BL21 (DE3). In the experiment, 0.65 mg of hGM-CSF was isolated from a 0.5 L flask culture of these E. coli and showed a 98% purity by SDS-PAGE analysis and silver staining. The bioactivity of this purified hGM-CSF was measured at an EC₅₀ of 16.4 ± 2 pM by a CCK8 assay in TF-1 human erythroleukemia cells.     

An Engineered Biliverdin-Compatible Cyanobacteriochrome Enables a Unique Ultrafast Reversible Photoswitching Pathway

Cyanobacteriochromes (CBCRs) are promising optogenetic tools for their diverse absorption properties with a single compact cofactor-binding domain. We previously uncovered the ultrafast reversible photoswitching dynamics of a red/green photoreceptor AnPixJg2, which binds phycocyanobilin (PCB) that is unavailable in mammalian cells. Biliverdin (BV) is a mammalian cofactor with a similar structure to PCB but exhibits redder absorption. To improve the AnPixJg2 feasibility in mammalian applications, AnPixJg2_BV4 with only four mutations has been engineered to incorporate BV. Herein, we implemented femtosecond transient absorption (fs-TA) and ground state femtosecond stimulated Raman spectroscopy (GS-FSRS) to uncover transient electronic dynamics on molecular time scales and key structural motions responsible for the photoconversion of AnPixJg2_BV4 with PCB (Bpcb) and BV (Bbv) cofactors in comparison with the parent AnPixJg2 (Apcb). Bpcb adopts the same photoconversion scheme as Apcb, while BV4 mutations create a less bulky environment around the cofactor D ring that promotes a faster twist. The engineered Bbv employs a reversible clockwise/counterclockwise photoswitching that requires a two-step twist on ~5 and 35 picosecond (ps) time scales. The primary forward P_fr → P_o transition displays equal amplitude weights between the two processes before reaching a conical intersection. In contrast, the primary reverse P_o → P_fr transition shows a 2:1 weight ratio of the ~35 ps over 5 ps component, implying notable changes to the D-ring-twisting pathway. Moreover, we performed pre-resonance GS-FSRS and quantum calculations to identify the Bbv vibrational marker bands at ~659,797, and 1225 cm⁻¹. These modes reveal a stronger H-bonding network around the BV cofactor A ring with BV4 mutations, corroborating the D-ring-dominant reversible photoswitching pathway in the excited state. Implementation of BV4 mutations in other PCB-binding GAF domains like AnPixJg4, AM1_1870g3, and NpF2164g5 could promote similar efficient reversible photoswitching for more directional bioimaging and optogenetic applications, and inspire other bioengineering advances.     

鞍钢鲅鱼圈4038m3高炉高比例使用中块焦炭生产实践

针对鞍钢股份有限公司鲅鱼圈钢铁分公司4038 m3高炉高比例使用中块焦炭后出现的风量萎缩、顺行变差,尤其是高炉休送风恢复过程中炉缸活跃性差、煤气流中心通道打不开、频繁崩滑料现象,提出采取合理匹配上下部调剂、组织好炉前渣铁排放、小焦布料制度以及改善休风料结构等措施.实践后,高炉实现了中块焦比为90 kg/t情况下的长期稳定顺行,休送风恢复未出现超时现象,适应了高比例使用中块焦炭的生产.     

X80管线钢埋弧焊匹配焊丝试验

通过分析高强度低合金钢焊缝熔敷金属常见显微组织对其力学性能的影响,确定X80管线钢用埋弧焊丝熔敷金属组织应以大量针状铁素体(AF)和少量粒状贝氏体(GB)的复合组织.从相变动力学原理出发,结合针状铁素体(AF)非自发形核机制和微合金组织韧化理论,选择Mn-Ni-Mo-Ti-B合金系进行X80管线钢匹配焊丝的试制.结果表明,合理选择和控制合金元素,可以获得理想的焊缝熔敷金属组织和强韧性以及低温韧性要求,试制的1号焊丝能够满足X80管线钢的使用要求.     

多传感器数据融合技术的现状及展望

阐述了多传感器数据融合的原理，对数据融合算法的研究动态进行了评述，指出了数据融合技术存在的问题，对今后的发展趋势进行了展望。     

Stage-efficiency of centrifugal extractor used in nuclear industry

1　INTRODUCTIONComparedwithamixer settlerorpulsedcolumnusedinthenuclearfuelreprocessingextraction pro cesses,ashort residence timecentrifugalextractorcatchesresearcher seyesbecauseofthe goodphaseseparationandasmallhold upvolume .Itshowsthefollowingadvantagesinthenuclearindustry[1] :1)Reducingsolventdegradationduetoradiation ;2 )Reducinginventoryofaqueousandorganicphases ,whichgivesrapidstart upandshut down ;3)Com pactdesignthatcontributestothereductionofplantcost .Manydifferenttypesofcen…     

Synthesis and characterization of novel aromatic condensation polymers containing rigid benzoxazole pendent groups

A new aromatic diamine monomer containing benzoxazole substituents was prepared by a multistep synthesis starting from 1,4‐dibromo‐2,5‐difluorobenzene. The diamine was polymerized with commercial aromatic dianhydride or dicarboxylic acid chloride monomers to provide several different poly(amic acid)s and polyamides with their inherent viscosities in the range of 0.24–0.46 dL/g. Thermal properties of these polymers including thermal imidization of poly(amic acid)s into polyimides were investigated by using FTIR, DSC, and TGA. © 2004 Wiley Periodicals, Inc. J Appl Polym Sci 92: 178–185, 2004     

对苯二甲酸加氢精制的反应规律

在高压磁力搅拌间歇反应釜中研究了粗对苯二甲酸在 Pd- C催化剂上加氢精制反应的规律。实验温度范围为 2 75～ 2 95℃ ,实验压力范围为 6 .0 8～ 8.0 8MPa(表压 )。研究发现加氢反应速率随着催化剂量和氢分压的增加而提高 ,在实验温度范围内 ,提高加氢温度 ,反应速率反而有所下降 ;投料中对苯二甲酸的质量浓度对反应速率也有一定影响 ,在 2 82℃反应条件下合适的质量分数为 2 5 %。     

设计企业项目管理信息系统的实现

结合国家“十五”重点科技攻关计划“城市规划、建设、管理与服务数字化工程”项目,天津水泥工业设计研究院承担了其中的“勘察设计企业信息化应用软件示范应用”专题,并获批立项,签署了专题任务书。两年来,院项目实施小组和软件公司进行了长期、紧密的合作,遵照标准化、系统化和集成化的原则,共同搭建了“工程协同设计管理平台”。在软件功能上,实现了设计企业管理信息系统的主要功能;在软件技术上,采用了目前最流行的软件技术,结合现代项目管理方法,为设计企业提供了组态灵活,方便实用,具有良好扩展性及开放性的软件产品。目前,该系统已经在部分工程项目上全面运行,取得了预期的效果。     

Ion-enrichment and ion-depletion of nanochannels based on electrochemical potential in a micro-nanofluidic chip

In this paper, we present a new approach for ion-enrichment and ion-depletion effect (IEID) in micro-nanofluidic chips without external power source. The method utilizes different reducibility of various electrodes in the weak oxidizing solution to generate the electrochemical potential and then induce IEID at a micro-nano junction. The results show that the average gray values of the micro-nano junction based on Al–Pt, Fe–Pt, and Cu–Pt electrodes increase from 14.7 to 40.2, 27.1, 15.0 after 20 s, and electric currents for Al–Pt, Fe–Pt, and Cu–Pt electrodes are 5.0, 2.9 and 0 nA respectively. Metal cations generating from oxidation–reduction reaction and electroosmotic convection flow are evaluated and their influences to IEID can be neglected in this case. The IEID method based on the electrochemical potential is power-free and weak flow convective that will be beneficial to the integrity of micro-nanofluidic chips and stability of IEID.