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Construction of shuttle, expression vector of human tumor necrosis factor alpha (hTNF-α) gene and its expression in a cyanobacterium, Anabaena sp. PCC 7120

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Science China(Life Sciences) 1999年第1期42卷 25-33页

作者：刘凤龙施定基商之狄邵宁徐旭东钟泽璞张宏斌吴锦银王捷江悦华赵树进林晨张雪艳吴旻彭国宏张海霞曾呈奎 1. Laboratory of Photosynthesis Institute of Botany Chinese Academy of Sciences 100093 Beijing China 2. The Guangzhou General Hospital of Guangzhou Military Command Guangzhou 510010 China 3. State Key Laboratory of Molecular Oncology Institute of Oncology Chinese Academy of Medicine 100021 Beijing China 4. EMBL Institute of Oceanology Chinese Academy of Sciences 266071 Qingdao China

The construction of the shuttle, expression vector of human tumor necrosis factor alpha (hTNF-a) gene and its expression in a cyanobacterium Anabaena sp. PCC 7120 was reported. The 700-bp hTNF cDNA fragments have been recovered from plasmid pRL-rhTNF, then inserted downstream of the promoter PpsbA in the plasmid pRL439. The resultant intermediary plasmid pRL-TC has further been combined with the shuttle vector pDC-8 to get the shuttle, expression vector pDC-TNF. The expression of the rhTNF gene in Escherichia coli has been analyzed by SDS-PAGE and thin-layer scanning, and the results show that the expressed TNF protein with these two vectors is 16.9 percent (pRL-TC) and 15.0 percent (pDC-TNF) of the total proteins in the cells, respectively, while the expression level of TNF gene in plasmid pRL-rhTNF is only 11.8 percent. Combined with the participation of the conjugal and helper plasmids, pDC-TNF has been introduced into Anabaena sp PCC 7120 by triparental conjugative transfer, and the stable transgenic strains have been obtained. The existence of the introduced plasmid pDC-TNF in recom-binant cyanobacterial cells has been demonstrated by the results of the agarose electrophoresis with the extracted plasmid samples and Southern blotting with α-32P labeled hTNF cDNA probes, while the expression of the hTNF gene in Anabaena sp. PCC 7120 has been confirmed by the results of Western blotting with extracted protein samples and human TNF-alpha monoclonal antibodies. The cytotoxicity.assays using the mouse cancer cell line L929 proved the cyto-toxicity of the TNF in the crude extracts from the transgenic cyanobacterium Anabaena sp. PCC 7120.

关键词： human tumor necrosis factor alpha (hTNF-α) Anabaena sp. PCC 7120 shuttle expression vector triparental conjugative transfer Southern and Western blottings cytotoxicity.

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Recombinant scorpion insectotoxin AaIT kills specifically insect cells but not human cellss

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Cell Research 2002年第2期12卷 143-150页

作者： SHENG JIAN JI, FENG LIU, ER Qiu LI, Yu XIAN ZHUThe National Laboratory of Protein Engineering and Plant Genetic Engineering, College of Life Sciences, Peking University, Beijing 100871, China 1The National Laboratory of Protein Engineering and Plant Genetic Engineering College of Life Sciences Peking University Beijing 100871 China 1The National Laboratory of Protein Engineering and Plant Genetic Engineering College of Life Sciences Peking University Beijing 100871 China 1The National Laboratory of Protein Engineering and Plant Genetic Engineering College of Life Sciences Peking University Beijing 100871 China 1The National Laboratory of Protein Engineering and Plant Genetic Engineering College of Life Sciences Peking University Beijing 100871 China

The nucleotide sequence deduced from the amino acid sequence of the scorpion insectotoxin AaIT was chemically synthesized and was expressed in Escherichia coli. The authenticity of this in vitro expressed peptide was confirmed by N-terminal peptide sequencing. Two groups of bioassays, artificial diet incorporation assay and contact insecticidal effect assay, were carried out separately to verify the toxicity of this recombinant toxin. At the end of a 24 h experimental period, more than 60% of the testing diamondback moth （Plutella xylostella） larvae were killed in both groups with LC50 value of 18.4 μM and 0.70 μM respectively. cytotoxicity.assay using cultured Sf9 insect cells and MCF-7 human cells demonstrated that the toxin AaIT had specific toxicity against insect cells but not human cells. Only 0.13 μM recombinant toxin was needed to kill 50% of cultured insect cells while as much as 1.3 μM toxin had absolutely no effect on human cells. Insect cells produced obvious intrusions from their plasma

关键词： Scorpion toxin AaIT prokaryotic expression cytotoxicity.

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Synthesis of 7a-Aza-benzo[fg]naphthacen-7-one

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Chinese Chemical Letters 2004年第3期15卷 269-272页

作者： ChengGangZHANG BoGangLI XiangGeZHOU GuoLinZHANG ChengduInstituteofOrganicChemistry ChineseAcademyofSciencesChengdu610041 ChengduInstituteofBiology ChineseAcademyofSciencesChengdu610041

2, 3, 11, 12-Tetramethoxy-3b, 4, 5, 6-tetrahydro-8H-7a-aza-benzo[fg]naphthacen-7-one 7 was synthesized via N-acyliminium ion cyclization of 6-hydroxy-1-(2, 3, 6, 7-tetramethoxy- phenanthren-9-yl-methyl)-piperidin-2-one 6 catalyzed by BF3OEt2. The stereostructure of the intermediate, 1-(2, 3, 6, 7-tetramethoxy-phenanthren-9-yl-methyl)-piperidine-2, 6-dione 5, was confirmed by X-ray crystallographic analysis. Compound 7 showed no cytotoxicity.

关键词： N-Acyliminium ion cyclization cytotoxicity.

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Enantiomeric Sesquiterpene Lactones from Senecio tsoongianus

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Chinese Chemical Letters 2002年第8期13卷 754-757页

作者： YuZHAO SimonPARSONS 等 DepartmentofTraditionalChineseMedicineandNaturalDrugResearch CollegeofPharmaceuticalSciencesZhejiangUniversityHangzhou310031 DepartmentofChemistry TheUniversityofEdinburghWestMa

From Senecio tsoongianus, two pairs of enantiomeric isomers, tsoongianolides A (1) and B (2), tsoongianolides C (3) and D (4) were isolated. Their structures were elucidated by 1D and 2D-NMR techniques and X-ray diffr... 详细信息

关键词： Sesquiterpenes eremophilanes X-ray cytotoxicity.

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Synthesis and in Vitro cytotoxicity.of a Novel Conjugate of Polyamide and Phospholipid

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Chinese Chemical Letters 2006年第5期17卷 599-602页

作者： Yi LIU Yu Lin DENG Rong ZHENG Pei Long ZHANG Hui ZHAI Kc Zhong LIU Chang Jin ZHU Department of Applied Chemistry Beijing Institute of Technology Beijing 100081 School of Life Science and Biotechnology Beijing Institute of Technology Beijing 100081

In order to increase the cell penetration and decrease the cytotoxicity.of the synthetic molecule, we designed and synthesized a DNA-binding ligand, which was a conjugate of polyamide and phospholipid. We examined the differences in the effect on cytotoxicity.between the conjugate and the native polyamide. As expected that the in vitro cytotoxicity.of the conjugate was obviously lower than that of the native polyamide either in HeLa cells or SH-SY5Y neuroblastoma cells.

关键词： Polyamide,phospholipid conjugate cytotoxicity.

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Synthesis and cytotoxicity.Evaluation of Some Isoquinoline Derivatives Related to 1-Arylnaphthalene Lignans

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Chinese Chemical Letters 2005年第10期16卷 1279-1282页

作者： Hong Xia DING Wei LU Chang Xin ZHOU Hai Bo LI Lei Xiang YANG Qi Jun ZHANG Xiu Mei WU Olivier BAUDOIN Jun Chao CAI Francoise GUERITTE Yu ZHAO Department of Traditional Chinese Medicine and Natural Drug Research College of Pharmaceutical Sciences Zhejiang University Hangzhou 31003 Shanghai Institute of Material Medica Chinese Academy of Sciences Shanghai 201203 Zhejiang Hisun Naturelite Pharmaceutical R＆D Co. Hangzhou 310006 Institute Chimie des Substances Naturelles CNRS AvenuedelaTerrasse 91198 Gif-sur-Yvette France

Two series of novel compounds designed as hybrids of 1-arylnaphthalene lignans with dihydroisoquinolines or isoquinolines were synthesized and evaluated for their cytotoxicities on human tumor cell lines, such as A549, Hela, PC-3 and KB. Some of the synthetic compounds exhibited their IC50 values on selected cell lines at μmol/L scale.

关键词： Isoquinoline derivatives 1-arylnaphthalene lignans synthesis cytotoxicity.

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Preparation and cytotoxicity.of Novel Aliphatic Polycarbonate Synthesized from Dihydroxyacetone

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Chinese Chemical Letters 2005年第5期16卷 572-574页

作者： LianShengWANG XueSongJIANG HaoWANG SiXueCHENG RenXiZHUO KeyLaboratoryofBiomedicalPolymersofMinistryofEducation DepartmentofChemistryWuhanUniversityWuhan430072 KeyLaboratoryofBiomedicalPolymersofMinistryofEducation DepartmentofChemistryWuhanUniversityWuhan430072／／DepartmentofChemistryXiaoganUniversityXiaogan432000

A new cyclic carbonate, 2,2-ethylenedioxypropane-1,3-diol carbonate (EOPDC), was synthesized through a two-step reaction from dihydroxyacetone dimer, and polymerized in bulk initiated by Sn(Oct)2 to give a high molecular weight polycarbonate. The structure of monomer and the polymer were characterized by FT-IR, 1H NMR, 13C NMR. The cytotoxicity.of the obtained polycarbonate was investigated by MTT assay.

关键词： Ring-opening polymerization aliphatic polycarbonate cytotoxicity.

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Saluenolide A,A Novel Eremophilanolide from Senecio saluenensis

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Chinese Chemical Letters 2002年第4期13卷 333-334页

作者： Yu ZHAO Peng WANG Simon PARSONS David W.H. RANKIN DepartmentofNaturalDrugResearch CollegeofPharmaceuticalSciencesZhejiangUniversityHangzhou310031

From Senecio saluenensis, a novel eremophilanolide Saluenolide A was isolated. Its structure was elucidated by 2D-NMR technique and X-ray diffraction.

关键词： Sesquiterpene eremophilanolide Senecio X-ray cytotoxicity.

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Differential recognition of MHC class I molecules of xeno-/allo-endothelial cells by human NK cells

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Science China(Life Sciences) 2000年第2期43卷 176-182页

作者：冯志民张晓峰王宏芳丰美福 1. State Key Lab of Biomembrane and Membrane Biotechnology Institute of Zoology Chinese Academy of Sciences 100080 Beijing China

Using human umbilical vein endothelial cells (HUVEC) and porcine aortic endothelial cells (PAEC) as target cells, human peripheral blood NK cells (PBNK) and NK92 cells as effector cells, the differential cytotoxicities of NK cells to allo- and xeno-endothelial cells were studied. The influence of MHC class I molecules on the cytotoxicity.of human NK cells was assayed using acid treatment, and blockades of MHC class I antigens, CD94 and KIR (NKB1). The results indicated that the killing of PAEC by the two kinds of NK cells is higher than that of HUVEC. After acid-treatment, the cytotoxicity.of the two kinds of NK cells to PAEC and HUVEC is significantly enhanced, but the magnitude of the enhancement is different. The enhancement of NK killing to acid treated HUVEC is much greater than that to PAEC. Blockade of CD94 mAb did not alter the NK cytotoxicity. while blockade of NKB1 mAb enhanced the cytotoxicity.of PBNK to HUVEC and PAEC by 95% and 29% respectively. The results above suggested that the differential recognition of MHC I molecules of xeno-endothelial cells by human NK cells could be the major reason for higher NK cytotoxicity.to PAEC. KIR might be the primary molecule that transduced inhibitory signals when endothelial cells were injured by NK cells.

关键词： human umbilical vein endothelial cells (HUVEC) porcine aortic endothelial cells (PAEC) peripheral blood natural killer cells (PBNK) NK92 acid treatment MHC class I molecules cytotoxicity.

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A New Cytotoxic Sterol Produced by an Endophytic Fungus from Castaniopsis fissa at the South China Sea Coast

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Chinese Chemical Letters 2004年第4期15卷 419-422页

作者： HouJinLI YongChengLIN L.L.P.VRIJMOED E.B.G.JONES DepartmentofAppliedChemistry ZhongshanUniversityGuangzhou510275 DepartmentofBiologyandChemistry CityUniversityofHongKongHongKong

A new sterol, ergosta-8(9),22-diene-3,5,6,7-tetraol(3β,5α,6β,7α,22E)(A) together with three known sterols: 3β, 5α,6β-trihydroxyergosta-7,22-diene (B), 3β-hydroxy-5α,8α- epidioxyer-gosta-6,22-diene (C) and ergosterol (D) were isolated from the mycelia of an unidentified endophytic fungus separated from Castaniopsis fissa (chestnut tree). Compound A exhibited potent selective cytotoxicity.against Bel-7402, NCI4460 and L-02 cell lines with IC50 values 8.445, 5.03, 13.621 μg/mL, respectively.

关键词： Endophytic fungus sterols ergosta-8(9) 22-diene-3,5,6,7-tetraol (3β,5α,6β,7α,22E) cytotoxicity.

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