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李春, Ying-Jin Yuan*, Chun Li, Zong-Ding Hu, Jin-Chuan Wu
Enzyme and Microbial Technology 29(2001)372~379,-0001,():
-1年11月30日
Signal transduction pathway for oxidative burst and taxol production was studied in suspension cultures of Taxus chinensis var. mairei induced by oligosaccharide from Fusarium oxysprum. Suspension cells exhibited an oxidative burst approximately 4h after challenge with oligosaccharide. The secondary metabolism including accumulation of extracellular phenolics and taxol production were enhanced, while the alkalization of the outer medium and activity of phenylalanine ammonia lyase (PAL) was promoted significantly. Pretreatment with the tyrosine (Tyr) kinase inhibitor herbimycin A abolished the oxidative burst and the subsequent induction of taxol production within 15 min after addition of elicitor. Suramin, a G-protein inhibitor, inhibited the oxidative burst if added up to 30 min after oligosaccharide treatment. The oxidative burst was also induced by the ion channel generator amphotericin B. In contrast, the ion channel blocker anthracene-9-carboxylate acid and Ca2 channel blocker nifedipine inhibited the oxidative burst within 45min after treatment. Oligosaccharide rapidly induced Tyr phosphrylation of receptor, which was inhibited by herbimycin A and anthracene-9-carboxylate acid. These responses were also inhibited by phospholipase C-neomycin. The response time of oligosaccharide to phospholipase C pathway was ca at 60 min. These data suggested that the activation of ion channels and phospholipase C follow Tyr kinase and G-protein in the signal pathway leading to the oxidative burst.
Fungal oligosaccharide, Signal transduction, Oxidative burst, Taxol biosynthesis, Taxus chinesis var., mairei
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李春, , 元英进, *马忠海, 胡宗定, 孙安慈, 胡昌序
Acta Botanica Sinica 2002, 44 (5): 598~602,-0001,():
-1年11月30日
在真菌(Fusarium oxysporum f . vasinfectum (Atkinson)Snyder et Hansen)寡聚糖诱导悬浮培养南方红豆杉(Taxus chinensis(Pilger)Rehd. var. mairei (Lemee et Lévl.)Cheng et L. K. Fu)细胞生产紫杉醇的体系中发现细胞出现凋亡,次生代谢增强。电镜观察到细胞核质和原生质出现凝集现象,液泡内出现大量的高电子致密体。核DNA经琼脂糖凝胶电泳,呈200 bp的整数倍的梯状条带(ladders);而对照组细胞核DNA 完整,呈大片段,细胞完整,细胞器发达,但紫杉醇合成速率很低。加入寡聚糖后,细胞防御系统开启,细胞生长停止,次生代谢物酚类物质大量积累且次生壁加厚,多酚氧化酶活性迅速提高,苯丙烷类代谢途径的关键酶苯丙氨酸解氨酶的活性在1h后急速提高,目的产物紫杉醇在诱导后72h达到峰值,比对照组提高了6倍,且细胞凋亡的出现与紫杉醇合成的峰值具有时间上的一致性。
真菌寡聚糖, 南方红豆杉, 凋亡, 次生代谢, 紫杉醇
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李春, Ying-Jin Yuan a, *, Chun Li a, Zong-Ding Hu a, Jin-Chuan Wu a, An-Ping Zeng b
Process Biochemistry 38(2002)193~198,-0001,():
-1年11月30日
A crude extract from Fusarium oxysprum caused significant cell apoptosis in suspension cultures of Taxus chinensis var. mairei as determined by transmission electron microscopy, DNA gel electrophoresis and terminal deoxynucleotidyl transferase-mediated dUTP nick labeling (TUNEL) analysis. Cells of T. chinensis var. mairei responded to the fungal elicitors by altering the general phenylpropanoid pathway and affecting the biosynthesis of taxol. The activity of L-phenylalanine ammonia lyase (PAL) increased appreciably within the first 36h of elicitor treatment, and then decreased rapidly. A large quantity of phenolic compounds accumulated in the culture medium. The maximum concentration of taxol was three times higher than that of the control. Cell apoptosis and taxol production are closely relevant in elicitor-treated culture of T. chinensis var. mairei .
Apoptosis, Taxol, Fungal elicitation, Suspension culture, Taxus chinensis var., mairei
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李春, SHUCHEN LIU, , * CHUN LI, LANGYI XIE, AND ZHU'AN CAO
JOURNAL OF BIOSCIENCE AND BIOENGINEERING Vol. 96, No.4, 349-353. 2003,-0001,():
-1年11月30日
The intracellular pH (pHi) of a,dicarboxylic acid producing Candida tropicalis was determined by a fluorescence technique using the pH-sensitive fluorescent probe 5(6)-carboxyfluorescein diacetate. Fermentations with n-tridecane substrate to produce a,-tridecanedioic acid were carried out in a 5-l bioreactor in which growth and production were separated. During the growth phase, the measured pHi values were varied from 5.65 to 6.15 in all the experiments performed under different constant pH-operating conditions. The specific rates of growth, sucrose consumption, CO2 production, and O2 consumption were correlated with pHi. Cytochrome P450 monooxygenase (P450), which catalyzes n-alkane hydroxylation, was only slightly expressed during the growth phase. During the first 6h of the production phase, P450 activity was induced rapidly accompanying higher pHi. A much higher level of P450 activity was observed at pHi of 6.55
intracellular pH,, Candida tropicalis,, tridecanedioic acid,, metabolic activity,, P450]
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李春, Chun Li, Ying-Jin Yuan*, Jin-Chuan Wu & Zong-Ding Hu
Biotechnology Letters 25: 1335~1343, 2003.,-0001,():
-1年11月30日
A structured kinetic model was established to describe the process of Taxol formation in suspension cultures of Taxus chinesis var. mairei induced by an oligosaccharide from Fusarium oxysporum. In this model, the role of intracellular starch as a storage carbon source had to be taken into account. Substrate uptake, culture growth, cell respiration, and secondary metabolites, predicted by the model, agreed with those obtained experimentally. The effective factors of oligosaccharide elicitation, ηe,j, defined as the ratio of the parameter values in the system with oligosaccharide to those in control, reflected the effects of the oligosaccharide on cell growth and Taxol production.
oligosaccharide,, plant cell culture,, structured kinetic model,, Taxol,, Taxus chinesis var., mairei
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