安成才
从事植物分子生物学及植物基因工程研究
个性化签名
- 姓名:安成才
- 目前身份:
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- 学位:
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学术头衔:
博士生导师
- 职称:-
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学科领域:
生物化学
- 研究兴趣:从事植物分子生物学及植物基因工程研究
现任北京大学生命科学学院教授,博士生导师,从事植物分子生物学及植物基因工程研究。1986年至1993年先后在日本三重大学、日本岗山大学分别获得硕士、博士学位和博士后研究,1993年北京大学生物系博士后研究,1995年出站后留校工作至今。1995年至1998年英国De Montfort大学兼职高级研究员,1997年至1998年英国牛津大学访问教授。现任国家出口管制专家支持体系的专家(聘书证号:03181);英国国际科学中心国际合作委员会顾问(No.:IC3001CN);世界发明家协会理事;美国帕拉丁国际科学中心客座研究员。主要工作:1)获得了抗病毒、抗肿瘤中药天花粉蛋白(TCS)基因及其突变体共12个,发现了天花粉蛋白诱导绒癌细胞、白血病细胞等肿瘤细胞凋亡新功能。找到TCS分子中与免疫反应重要相关氨基酸位点,特别是与肿瘤细胞识别与入胞作用相关的关键氨基酸残基,并证明了TCS蛋白中与靶细胞受体相互作用的入胞机制,为消除或改善药物蛋白的免疫原性及其毒副作用,为实现肝癌、乳腺癌等靶细胞专一性杀伤导向药物的研发奠定了重要基础。2)克隆出与水稻矮化/矮化调控重要相关的水稻赤霉素20氧化酶基因(rga5)及其同工酶基因(rsd-1),阐明了该基因在赤霉素合成代谢、以及水稻生长发育中的重要调节功能,培育出促进植株生长或植株矮化的转基因水稻,并创建了T-Linker PCR新方法。3)从蓝藻水华中分离出具有显著溶藻效果的溶藻菌、噬藻体和食藻原生动物等溶藻体,完成了SP4噬藻体的全基因组测序分析,其中对SP4噬藻体的宿主决定性相关因子及其侵染藻细胞分子机制进行研究。,并建立了蓝藻水花的生物控制技术体系。4)克隆和鉴定病原物诱导型植物防卫基因(pal、CHS)及其启动子,确定其顺式作用元件及其控制的基因表达模式;鉴定和分离植物防卫基因的反式因子及其编码基因,研究其结构及其与顺式因子的作用特性; 反式因子基因的植物/细胞转化,探讨反式因子对防卫基因(pal、CHS)及其防卫反应的调控机理。
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2833
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0
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成果阅读
505
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成果数
9
【期刊论文】Expression and puriWcation of Huwentoxin-I in baculovirus system
安成才, Wenjie Jia, Xinyue Zhanga, Huicong Hua, Jiyuan Chena, Yin Gaoa, Songping Liangb, Chengcai Ana, *
Protein Expression and PuriWcation 41(2005)454-458,-0001,():
-1年11月30日
Huwentoxin-I (HWTX-I) is a novel neurotoxin isolated from the venom of Orinithoctonus huwena. Based on its biological activity, HWTX-I could be developed as a pain-killer for clinical purpose. Production of HWTX-I by the bacterium or yeast expression systems resulted in poor yields and the puriWed protein was proved to have lower biological activity than that of native one. So, for the Wrst time, we introduced a new method to express HWTX-I gene in Sf9 cells using baculovirus expression system. Recombinant HWTX-I was recognized by Western blotting and then puriWed by nickel-chelating aYnity chromatography under native conditions. Recombinant HWTX-I showed identical amino acid sequence as native form and exhibited similar eVect on muscular transmission with that of native form. These results indicate that the baculovirus expression system and native puriWcation strategy are viable ways to produce active HWTX-I.
Huwentoxin-I, Nickel-chelating aYnity chromatography, Baculovirus expression system
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安成才, Shuang-Li Mi, Cheng-Cai An*, Ye Wang, Ji-Yuan Chen, Nan-Ying Che, Yin Gao, Zhang-Liang Chen
Archives of Biochemistry and Biophysics 434(2005)258-265,-0001,():
-1年11月30日
Trichomislin, a novel ribosome-inactivating protein, was cloned from the genome of Trichosanthes kirilowii Maxim. The gene was recombined to prokaryotic expression vector and the protein was puriWed by cation-exchange chromatography. The secondary structure of trichomislin was measured by circular-dichroism analysis and the ratios of α-helices and β-sheets were calculated. Trichomislin could inhibit the synthesis of protein in rabbit reticulocyte lysate systems and its reaction me hanism was to inactivate ribosome as an rRNA N-glycosidase. Antitumor analyses indicated trichomislin induced the apoptosis and inhibited the growth of choriocarcinoma cells. Further investigation showed that trichomislin could bind to and enter choriocarcinoma cells, and then increase the caspase-3 activity in a time-dependent manner. At the same time, the concentration of cytochrome c in cytosol increased while that in mitochondria decreased. These results suggested that trichomislin induced apoptosis by releasing cytochrome c from mitochondria which then triggered the caspase family member activation.
Ribosome-inactivating protein, Choriocarcinoma, Apoptosis, Mitochondria, Caspase
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【期刊论文】SiteFinding-PCR: a simple and efficient PCR method for chromosome walking
安成才, Guihong Tan, Yin Gao, Miao Shi, Xinyue Zhang, Shanping He, Zhangliang Chen and Chengcai An*
Nucleic Acids Research, 2005, Vol. 33, No.13 e122,-0001,():
-1年11月30日
In this paper, we present a novel PCR method, termed SiteFinding-PCR, for gene or chromosome walking. The PCR was primed by a SiteFinder at a low temperature, and then the target molecules were amplified exponentially with gene-specific and SiteFinder primers, and screened out by another gene-specific primerand a vector primer.However, non-target molecules could not be amplified exponentially owing to the suppression effect of stem-loop structure and could notbescreenedout. This simplemethodproved to be efficient, reliable, inexpensive and time-saving, andmay be suitable for themolecules for which genespecific primers are available. More importantly, large DNA fragments can be obtained easily using this method. To demonstrate the feasibility and efficiency of SiteFinding-PCR, we employed this method to do chromosomewalkingandobtained 16positive results from 17 samples.
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安成才, L. Wang, C. An, W. Qian, T. Liu, J. Li, Z. Chen
Plant Cell Rep (2004) 22: 513-518,-0001,():
-1年11月30日
A rice PAL (phenylalanine ammonia-lyase) gene sequence (rPAL-P5), which is highly similar to and likely the same as a previously described rice ZB8PAL gene, including the 50-upstream and exon I coding regions of PAL, was isolated using PCR amplification. The expression of several PALs, including rPAL-P5, was strongly induced following inoculation with Pyricularia oryzae or treatment with a P. oryzae elicitor. To identify the promoter region induced by the P. oryzae elicitor, we constructed and subsequently transformed rPAL-P5 promoter deletion series into rice calli using particle bombardment. Results from both elicitor-inducible reporter gene and gel mobility shift assays demonstrated that the sequence -349 to -256 of the rPAL-P5 promoter includes a cis-element involved in the induction of P. oryzae.
Phenylalanine ammonia-lyase, cis-Element, Elicitor, Rice calli, Promoter
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安成才, Ye Wang, Shuang-Li Mi, Mei-Yan Lou, Yin Gao, Zhang-Liang Chen and Cheng-Cai An
[Frontiers in Bioscience 10, 2279-2284, September 1, 2005],-0001,():
-1年11月30日
Trichosanthin (TCS) is a ribosome-inactivating protein (RIP) which can inhibit the growth of human choriocarcinoma (JAR) cells. There are no clear mechanisms to discover the interaction pathway and cytotoxicity of TCS in JAR cells. In this paper, we showed the distribution and transport of ndogenously expressed TCS in JAR cells. Enhanced Green Fluorescence Protein (EGFP), fused with TCS, was applied as a reporter to track the behavior of TCS in JAR cells. Firstly, we investigated the expression stability of EGFP and physiological effects on JAR cells. A stable cell line expressing EGFP was created, which could reproduce and express EGFP even if transplanted into nude mice. Based on the proved stability and feasibility of EGFP in cultured cells and in vivo, the fusion gene of EGFP and TCS was constructed and transfected into JAR cells by liposome. The fluorescence microscopy showed that TCS-EGFP fusion gene was expressed in JAR cells in 24 to 48 hours and the fluorescence spread in cytoplasm mainly and in nucleus partially, which could trace the distribution and transport of TCS-EGFP in JAR cells. Most of fluorescent cells died after 48 hours for the cytotoxicity of expressed TCS-EGFP. These results first reported a stable expression and tracing method by EGFP in JAR cells, and provided theoretical basis to apply TCS in cancer therapy.
trichosanthin,, choriocarcinoma cells,, EGFP,, liposome-induced transfection
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安成才, Yan Yuanxin, , An Chengcai, *, Li Li, Gu Jiayu, Tan Guihong and Chen Zhangliang
Nucleic Acids Research, 2003, Vol. 31, No.12 e68,-0001,():
-1年11月30日
Dozens of PCR-based methods are available for chromosome walking from a known sequence to an unknown region. These methods are of three types: inverse PCR, ligation-mediated PCR and randomly primed PCR. However, none of them has been generally applied for this purpose, because they are either difficult or inefficient. Here we describe a simple and efficient PCR strategy
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安成才, Chun-yang ZHANG*, Yi-xuan GONG†, Hui MA*, Cheng-cai AN† and Die-yan CHEN*
Biochem. J. (2001) 355, 653-661 (Printed in Great Britain),-0001,():
-1年11月30日
The type-I ribosome-inactivating protein trichosanthin (TCS) has a broad spectrum of biological and pharmacological activities, including abortifacient, anti-tumour and anti-HIV activities. We have found for the first time that TCS stimulated the production of reactive oxygen species (ROS) in JAR cells (a human choriocarcinoma cell line) in a time- and concentrationdependent manner by using the fluorescent probe 2',7'-dichlorofluorescein diacetate with confocal laser scanning microscopy. ESR spectral studies and the inhibition of ROS formation by the superoxide radical anion (O2 −d) scavenger superoxide dismutase, the H3O2 scavenger catalase and the hydroxyl radical (OHd) scavenger mannitol suggested the involvement of O2−d, H2O2 and OHd. TCS-induced ROS formation was shown to be dependent on the presence of both extracellular and intracellular Ca2+; moreover, ROS production paralleled the intracellular Ca2+ elevation induced by TCS, suggesting that ROS production might be a consequence of Ca2+ signalling. TCS-induced activation of caspase-3 was initiated within 2 h; however, TCS-induced production of ROS was initiated within 5min, suggesting that the production of ROS preceded the activation of caspase-3. Simultaneous observation of the nuclear morphological changes via two-photon laser scanning microscopy and ROS production via confocal laser scanning microscopy revealed that ROS is involved in the apoptosis of JAR cells. The involvement of ROS was also confirmed by the inhibition of TCS-induced cell death by the antioxidant Trolox and the ROS scavengers catalase and mannitol. Diethylenetriaminepenta-acetic acid, an inhibitor of metal-facilitated OHd formation, markedly inhibited TCS-induced cell death, suggesting that TCS induced OHd formation via the Fenton reaction. The finding that ROS is involved in the TCS-induced apoptosis of JAR cells might provide new insight into the antitumour and anti-HIV mechanism of TCS.
calcium,, caspase-3,, hydrogen peroxide,, hydroxyl radical,, superoxide radical anion.,
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【期刊论文】Capillary electrophoresis and circular dichroism study of trichosanthin and its mutants
安成才, Chun-yang Zhanga, b, Cheng-cai Ana, *, Rong-ying Wangb, Yi-xuan Gonga, Hui Mab, Die-yan Chenb, Zhang-liang Chena
Talanta 57(2002)467-473,-0001,():
-1年11月30日
The type-I ribosome-inactivating protein trichosanthin (TCS) has a broad spectrum of biological and pharmacological activities, including abortifacient, anti-tumor and anti-human-immunodeficiency-virus (anti-HIV). In this study, circular dichroism (CD) and capillary electrophoresis were used for the first time to study TCS and its two TCS mutants of Y55G TCS (tyrosine 55 converted to glycine) and FYY140-142GSA TCS (tripeptide phenylalanine-tyrosine-tyrosine 140-142 converted to glycine-serine-alanine). The results indicated that the substitution of amino acids changed the secondary structures and the hydrophobility of TCS. Moreover, both Y55G TCS and FYY140-142GSA TCS demonstrated attenuated cytotoxicity and reactive oxygen species (ROS) production in human choriocarcinoma cells (JAR cells) as compared to natural TCS and wild-type TCS. Our results demonstrated the cytotoxicity of TCS on JAR cells and TCS-induced production of ROS might be TCS-conformational related, suggesting that CD and capillary electrophoresis study might throw new insight into the anti-tumor and anti-HIV mechanism of TCS.
Circular dichroism, Trichosanthin, Capillary electrophoresis
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