Complementation analysis reveals a potential role of human ARV1 in GPI anchor biosynthesis

ARV1 is involved in regulating lipid homeostasis but also in the biosynthesis of glycosylphosphatidylinositol (GPI) in Saccharomyces cerevisiae. Here, we examined whether human ARV1 can complement the role of yeast ARV1 in GPI biosynthesis. Overexpression of human ARV1 could rescue the phenotypes associated with GPI anchor synthesis defect in the yeast arv1Δ mutant. The results suggest that Arv1 function in GPI biosynthesis may be conserved in all eukaryotes, from yeast to humans. Copyright © 2015 John Wiley & Sons, Ltd.     

板材FMS下位机人机接口的设计与实现

介绍板材ＦＭＳ下位机冲床单元人机接口的设计,操作界面简单明了,分类化显示,每个设备都有一个寒带的画面;将系统状态,操作设备的状态和操作对象的状态作关联化显示,同时提高了系统的安全可靠性。人机接口采用ＳＴＤ工控机,提高了系统配置的模块化和灵活性,该系统已在生产中成功运行多年。     

高危型人乳头瘤病毒检测作为宫颈癌前病变首筛项目的研究

目的探讨高危型人乳头瘤病毒（HPV）检测作为宫颈癌前病变首筛项目的应用价值。方法对1 013例慢性宫颈炎或疑似宫颈病变患者采用液基细胞学（LCT）检查及第二代杂交捕获技术（HC2）高危型HPV检测进行宫颈癌筛查,并与组织病理学比较。LCT检查以LCT≥非典型鳞状细胞（ASCUS）/非典型腺细胞（AGUS）为阳性,HC2检测以HC2≥1.0ng·L-1为阳性,任一项结果阳性者行阴道镜下取活检,并以宫颈活检病理结果为确诊金标准。结果 LCT检查对宫颈癌前病变敏感度、特异度、阳性预测值、阴性预测值分别为：89.61%、97.08%、27.27%、94.32%,HC2检测对宫颈癌前病变敏感度、特异度、阳性预测值、阴性预测值分别为：96.10%、70.03%、43.79%、98.67%;HC2检测的敏感度、阳性预测值及阴性预测值均显著高于LCT检查（P〈0.05）,但特异度显著低于LCT检查（P〈0.05）。LCT联合HC2检测（LCT、HC2任一阳性）阳性符合率为19.54%,HC2检测阳性符合率（阳性预测值）明显高于LCT联合HC2检测（P〈0.05）。结论在进行宫颈癌前病变筛查的流程中,将高危型HPV检测作为筛查流程的首筛项目,有助于降低宫颈癌前病变的漏诊率;高危型HPV检测阳性的患者再循流程行LCT,可提高宫颈癌前病变筛查的特异性;高危型HPV检测阴性的患者不必行进一步检查,有助于降低检查成本,避免不必要的浪费。     

工程设计的发展趋势

工程设计是以改造客观世界为目的的创造性劳动,它直接决定产品的质量和寿命,是工程的灵魂。在当前信息化与高科技的社会背景下,本文作者就其工程设计的发展趋势作了论述。     

基于模型的人体运动跟踪

在计算机视觉领域,人体运动分析的的研究正因其广泛的应用前景而越来越受到研究 重视,对图像序列中的人体运动进行了跟踪是其中的关键技术。由于人体运动的特殊复杂性,已有的研究方法都对人体加上了许多限制条件。     

人尿中活性物质的开发

本文叙述了近年来人尿中活性物质的开发状况,列举了从人尿中提取的11种生物药品及其应用,并提出今后发展的方向。     

Mutational analysis of structure--activity relationships in human tumor necrosis factor-alpha

To determine the region of human tumor necrosis factor-alpha(TNF-), essential for cytotoxic activity against mouse L-M cells,single amino-acid-substituted TNF- mutant proteins (muteins)were produced in Escherichia coli by protein engineering techniques.An expression plasmid for TNF- was mutagenized by passage throughan E.coli mutD5 mutator strain and by oligonucleotide-directedmutagenesis. Approximately 100 single amino-acid-substitutedTNF- muteins were produced and assayed for cytotoxic activity.The cytotoxic activities of purified TNF- muteins, e.g. TNF-31T,-32Y, -82D, -85H, -115L, -141Y, -144K and -146E, were < 1%of that of parent TNF-. These results indicate that the integrityof at least four distinct regions of the TNF- molecule is requiredfor full biological activity. These regions are designated asfollows: region I, from position 30 to 32; region II, from position82 to 89; region III, from position 115 to 117; region FV, fromposition 141 to 146. In addition, TNF-141Y could not completelycompete with parent TNF- for binding to the receptor. This demonstratesthat region IV, and at least aspartk acid at position 141, mustbe involved in the TNF receptor binding site.     

Efficient Total Chemical Synthesis of 13C=18O Isotopomers of Human Insulin for Isotope‐Edited FTIR

Isotope‐edited two‐dimensional Fourier transform infrared spectroscopy (2 D FTIR) can potentially provide a unique probe of protein structure and dynamics. However, general methods for the site‐specific incorporation of stable ¹³C=¹⁸O labels into the polypeptide backbone of the protein molecule have not yet been established. Here we describe, as a prototype for the incorporation of specific arrays of isotope labels, the total chemical synthesis—via a key ester insulin intermediate—of 97 % enriched [(1‐¹³C=¹⁸O)Phe^B24] human insulin: stable‐isotope labeled at a single backbone amide carbonyl. The amino acid sequence as well as the positions of the disulfide bonds and the correctly folded structure were unambiguously confirmed by the X‐ray crystal structure of the synthetic protein molecule. In vitro assays of the isotope labeled [(1‐¹³C=¹⁸O)Phe^B24] human insulin showed that it had full insulin receptor binding activity. Linear and 2 D IR spectra revealed a distinct red‐shifted amide I carbonyl band peak at 1595 cm^?1 resulting from the (1‐¹³C=¹⁸O)Phe^B24 backbone label. This work illustrates the utility of chemical synthesis to enable the application of advanced physical methods for the elucidation of the molecular basis of protein function.     

Novel Redox-Dependent Esterase Activity (EC 3.1.1.2) for DJ-1: Implications for Parkinson’s Disease

Mutations the in human DJ-1 (hDJ-1) gene are associated with early-onset autosomal recessive forms of Parkinson’s disease (PD). hDJ-1/parkinsonism associated deglycase (PARK7) is a cytoprotective multi-functional protein that contains a conserved cysteine-protease domain. Given that cysteine-proteases can act on both amide and ester substrates, we surmised that hDJ-1 possessed cysteine-mediated esterase activity. To test this hypothesis, hDJ-1 was overexpressed, purified and tested for activity towards 4-nitrophenyl acetate (pNPA) as µmol of pNPA hydrolyzed/min/mg·protein (U/mg protein). hDJ-1 showed maximum reaction velocity esterase activity (V_max = 235.10 ± 12.00 U/mg protein), with a sigmoidal fit (S_0.5 = 0.55 ± 0.040 mM) and apparent positive cooperativity (Hill coefficient of 2.05 ± 0.28). A PD-associated mutant of DJ-1 (M26I) lacked activity. Unlike its protease activity which is inactivated by reactive oxygen species (ROS), esterase activity of hDJ-1 is enhanced upon exposure to low concentrations of hydrogen peroxide (<10 µM) and plateaus at elevated concentrations (>100 µM) suggesting that its activity is resistant to oxidative stress. Esterase activity of DJ-1 requires oxidation of catalytic cysteines, as chemically protecting cysteines blocked its activity whereas an oxido-mimetic mutant of DJ-1 (C106D) exhibited robust esterase activity. Molecular docking studies suggest that C106 and L126 within its catalytic site interact with esterase substrates. Overall, our data show that hDJ-1 contains intrinsic redox-sensitive esterase activity that is abolished in a PD-associated mutant form of the hDJ-1 protein.     

MicroRNA-331-3p Suppresses Cervical Cancer Cell Proliferation and E6/E7 Expression by Targeting NRP2

Aberrant expression of microRNAs (miRNAs) is involved in the development and progression of various types of cancers. In this study, we investigated the role of miR-331-3p in cell proliferation and the expression of keratinocyte differentiation markers of uterine cervical cancer cells. Moreover, we evaluated whether neuropilin 2 (NRP2) are putative target molecules that regulate the human papillomavirus (HPV) related oncoproteins E6 and E7. Cell proliferation in the human cervical cancer cell lines SKG-II, HCS-2, and HeLa was assessed using the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt (MTS) assay. Cellular apoptosis was measured using the TdT-mediated dUTP nick end labeling (TUNEL) and Annexin V assays. Quantitative RT-PCR was used to measure the messenger RNA (mRNA) expression of the NRP2, E6, E7, p63, and involucrin (IVL) genes. A functional assay for cell growth was performed using cell cycle analyses. Overexpression of miR-331-3p inhibited cell proliferation, and induced G2/M phase arrest and apoptosis in SKG-II, HCS-2 and HeLa cells. The luciferase reporter assay of the NRP2 3′-untranslated region revealed the direct regulation of NRP2 by miR-331-3p. Gene expression analyses using quantitative RT-PCR in SKG-II, HCS-2, and HeLa cells overexpressing miR-331-3p or suppressing NRP2 revealed down-regulation of E6, E7, and p63 mRNA and up-regulation of IVL mRNA. Moreover, miR-331-3p overexpression was suppressed NRP2 expression in protein level. We showed that miR-331-3p and NRP2 were key effectors of cell proliferation by regulating the cell cycle, apoptosis. NRP-2 also regulates the expression of E6/E7 and keratinocyte differentiation markers. Our findings suggest that miR-331-3p has an important role in regulating cervical cancer cell proliferation, and that miR-331-3p may contribute to keratinocyte differentiation through NRP2 suppression. miR-331-3p and NRP2 may contribute to anti-cancer effects.