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Search Results: 1 - 10 of 39213 matches for " YAO Shan-jing "
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Study on Disulfide Bond Formation Protein A in Escherichia coli
大肠杆菌二硫键形成蛋白A(DsbA)研究进展

LUO Man,GUAN Yi-Xin,YAO Shan-Jing,
罗曼
,关怡新,姚善泾

生物工程学报 , 2007,
Abstract: Disulfide bond formation protein A, DsbA, is one of the important proteins located in E. coli periplasm, which is a foldase facilitating the folding of nascent secreted proteins, especially for those with many pairs of disulfide bonds. The crystal structure and phylogenetic analysis of DsbA and DsbA-mediated protein folding, alternatively in vivo and in vitro, are summarized. Both the extremely low pK_a of Cys~ 30 , about 3.5, and the destabilizing effect of the active site disulfide contribute to its strong oxidizing power. The Cys~ 30 is also considered as the most important residue closely related to its activity using site-directed mutagenesis methodology. DsbA could effectively assist proteins folding, both in vivo coexpressed with the target protein, and in vitro replenished as foldases. Moreover, DsbA also has the chaperone-like activity in the assistant refolding of genetically engineered inclusion bodies.
Study on Disulfide Bond Formation Protein A in Escherichia coli
大肠杆菌二硫键形成蛋白A(DsbA)研究进展

LUO Man,GUAN Yi-Xin,YAO Shan-Jing,
罗曼
,关怡新,姚善泾

微生物学报 , 2007,
Abstract: Disulfide bond formation protein A, DsbA, is one of the important proteins located in E. coli periplasm, which is a foldase facilitating the folding of nascent secreted proteins, especially for those with many pairs of disulfide bonds. The crystal structure and phylogenetic analysis of DsbA and DsbA-mediated protein folding, alternatively in vivo and in vitro, are summarized. Both the extremely low pKa of Cys30, about 3.5, and the destabilizing effect of the active site disulfide contribute to its strong oxidizing power. The Cys30is also considered as the most important residue closely related to its activity using site-directed mutagenesis methodology. DsbA could effectively assist proteins folding, both in vivo coexpressed with the target protein, and in vivo replenished as foldases. Moreover, DsbA also has the chaperone-like activity in the assistant refolding of genetically engineered inclusion bodies.
Formation of dimers in refolding of recombinant human interferon-γ by ion-exchange chromatography
离子交换层析复性重组人γ-干扰素折叠二聚体的形成

JIN Ting,GUAN Yi-xin,YAO Shan-jing,
靳挺
,关怡新,姚善泾

浙江大学学报(农业与生命科学版) , 2006,
Abstract: SP Sepharose Fast Flow离子交换层析柱作为蛋白质复性系统,采用尿素梯度法进行重组人γ-干扰素包涵体蛋白质的复性实验.结果表明,尿素梯度离子交换层析法能有效地复性重组人γ-干扰素包涵体,复性后的重组人γ-干扰素的纯度达95%,蛋白收率达54%,比活为7.5×105IU?mg-1.以Superdex 75凝胶作为体积排阻层析介质,对离子交换层析复性的样品进行分析,表明离子交换层析复性的样品中没有重组人γ-干扰素聚集体存在.荧光光谱分析表明重折叠的重组人γ-干扰素的构象接近于其天然二聚体构象.
Mechanism of Effects of Allyl Compounds on Asymmetric Reduction of Baker''''s Yeast
烯丙基化合物对面包酵母立体选择性的影响机理

LI Yan,NI Hong-liang,YAO Shan-jing,
李彦
,倪宏亮,姚善泾

过程工程学报 , 2007,
Abstract: 添加烯丙基化合物可以控制酵母催化不对称还原反应的立体选择性,以4-氯-3-羰基丁酸乙酯(COBE)为模型底物,用丙烯酰胺和烯丙基醇对酵母进行预处理,产物(S)-4-氯-3-羟基丁酸乙酯(S-CHBE)的对映体过量值(e.e.)可以增加至94%;而用烯丙基溴预处理,则可以得到R型产物.通过DNS法和API ZYM试验条证明烯丙基类化合物对细胞及胞内酶系存在毒害作用,且作用大小取决于烯丙基所带基团.预处理后的酵母细胞经PBS缓冲液多次洗涤后,其活性可以逐渐得到恢复.随着底物浓度的增加,经烯丙基醇预处理的酵母细胞,其S型产物的e.e.值维持在90%左右,而经烯丙基溴处理后,反应得到的R型产物的e.e.值则从90%下降到45%左右.说明烯丙基类化合物竞争性地吸附于酶的活性区域,且其所带基团决定了被吸附酶的类型.以抑制剂类似物处理酵母细胞,其反应结果说明酵母体内催化不对称还原反应的两类酶存在着空间位阻效应.
Effects of Acrylamide on the Activity and Structure of Human Brain Creatine Kinase
Qing Sheng,He-Chang Zou,Zhi-Rong Lü,Fei Zou,Yong-Doo Park,Yong-Bin Yan,Shan-Jing Yao
International Journal of Molecular Sciences , 2009, DOI: 10.3390/ijms10104210
Abstract: Acrylamide is widely used worldwide in industry and it can also be produced by the cooking and processing of foods. It is harmful to human beings, and human brain CK (HBCK) has been proposed to be one of the important targets of acrylamide. In this research, we studied the effects of acrylamide on HBCK activity, structure and the potential binding sites. Compared to CKs from rabbit, HBCK was fully inactivated at several-fold lower concentrations of acrylamide, and exhibited distinct properties upon acrylamide-induced inactivation and structural changes. The binding sites of acrylamide were located at the cleft between the N- and C-terminal domains of CK, and Glu232 was one of the key binding residues. The effects of acrylamide on CK were proposed to be isoenzyme- and species-specific, and the underlying molecular mechanisms were discussed.
A Colorimetric Method to Assay Biological Activity of Recombinant Human IFN-γ
重组人γ干扰素复性过程中体外活性检测方法研究*

FEI Zheng-Zheng GUAN Yi-Xin YAO Shan-Jing,
费峥峥
,关怡新,姚善泾

微生物学通报 , 2004,
Abstract: 结晶紫染色法由于具有客观、精确的特点,适用于大量抗肿瘤药物的药敏测定。本实验对此方法进行了探讨和条件优化,考察了结晶紫染液的用量及作用时间、溶媒提取时间、孔板效应及边缘效应等,并将其应用于重组人γ干扰素包涵体初步复性的体外活性检测之中。
Hydroxylation of 16α, 17α-epoxy-4-pregenene-3,20-dione by Absidia coerulea with Pseudo-crystallo Feed
拟结晶投料环氧黄体酮犁头霉羟化过程研究

WANG Jia,GUAN Yi-Xin,WANG Hai-Qing,YAO Shan-Jing,
王嘉
,关怡新,王海清,姚善泾

生物工程学报 , 2006,
Abstract: The 11-hydroxylation of 16alpha,17alpha-epoxy-4-pregenene-3,20-dione as a useful intermediate for the preparation of hormones can be achieved by the mycelium of Absidia coerulea at higher conversion rate than using other strains. In this paper 16alpha,17alpha-epoxy-4-pregenene-3,20-dione mixed with a little water, beta-cyclodextrin, Tween-80 was introduced into the fermentation broth after ultrasonication to increase pseudo-water-solubility of the hydrophobic substrate. This pseudo-crystallo feed could avoid the toxicity of organic solvents and was more available for the microbial transformation. The multi layer feed-forward neural network was used to setup a model which indicated the relationship between medium and feed components and the conversion rate. Particle swarm optimization (PSO), which was a stochastic global optimization algorithm and of which the convergence speed was high, was applied to obtain the optimal concentration of the medium and feed components. At optimum conditions with the pseudo-crystallo feed, the conversion rate of 16alpha,17alpha-epoxy-4-pregenene-3,20-dione at an initial concentration of 10 g/L was 87.5% in shaking flasks. The conversion rate of the substrate was up to 86.6% at higher concentration of 20 g/L feed in a 3.7 L fermentor.
Preliminary Proteome Analysis for Saccharomyces cerevisiae under Different Culturing Conditions
在不同碳源培养条件下酿酒酵母的蛋白质组解析

ZHANG Hui-Min YAO Shan-Jing,PENG Li-Feng Kazuyuki Shimizu,
张慧敏
,姚善泾,彭立凤,清水和幸

生物工程学报 , 2004,
Abstract: For the investigation of the metabolic regulation of Saccharomyces cerevisiae under different culture conditions, the proteins of cell utilizing various carbon sources were separated by two-dimensional electrophoresis with immobilized pH gradients as the first dimension and SDS-PAGE as the second. Samples were taken in the log phase of batch culture using glucose or lactic acid as carbon source, while another sample was taken from the broth when glucose was consumed up and ethanol accumulated in the previous phase was further metabolized. After electrophoresis, the protein spots were detected by silver-stain in a Hoefer Automated Gel Stainer with a protein silver staining kit. Silver-stained gels were scanned and digitized to create computer images. About 500 protein spots were detected by employing the 2D proteome image analysis system Image Master 2D Elite and SWISS-2DPAGE proteome database. Most of the protein expressed and involved in the glycolysis, pentose phosphate (PP) pathway, anaplerotic pathway, as well as TCA cycle were analyzed. The metabolism regulation of protein level for Saccharomyces cerevisiae under various carbon sources, as well as during different phase of growth, was studied. The expression of several glycolytic en~zymes(glk, pgi, pgk, eno, pyk) was up-regulated while the expression of enzymes in oxidative pentose phosphate pathway(zwf, gnd) was down-regulated when ethanol and lactic acid were taken as carbon source. Simultaneously, frucotose1,6-biphosphatase was found to be up-regulated due to the gluconeogenic requirement. Citrate synthase and Malate dehydrogenase do not exhibit significant difference, indicating TCA cycle is necessary when utilizing glucose, ethanol or lactic acid as carbon source. Thus, the NADPH loss due to the repressed pentose phosphate pathway could be compensated by TCA cycle in cases of ethanol and lactic acid. The expression of malic enzyme and isocitrate lyase are activated to a large extent when metabolizing ethanol, indicating glyoxylate shunt is essential in transferring ethanol to generate four carbon precursors for the biosynthesis and the NADP-dependent malic enzyme could also serve as compensation mechanism for NADPH loss in this case.
Soluble expression of recombinant cyclophilin A in Escherichia coli
重组亲环蛋白A的可溶性表达

YAN Xiao,XU Li-ren,GUAN Yi-xin,YAO Shan-jing,
闫啸
,徐立仁,关怡新,姚善泾

浙江大学学报(农业与生命科学版) , 2010,
Abstract: 为获得高表达量的可溶亲环蛋白A(cyclophilin A,CypA),对重组CypA质粒表达和工程菌的培养过程进行研究.首先将pRSET-CypA质粒转入Escherichia coli BL21菌株,获得表达CypA的重组工程菌;然后以2×YT为基本培养基,对培养基组成包括碳源、氮源和氨苄青霉素浓度进行优化,并考察接种量和诱导条件(诱导剂浓度、诱导时机、诱导后培养时间)等对目标蛋白表达量的影响.结果表明:通过控制培养温度和转速可大大减少CypA包涵体的量;以甘露醇为碳源能显著提高目标蛋白的表达量;优化的培养基组成为蛋白胨16 g·L-1,酵母提取物10 g·L-1,NaCl 5 g·L-1,甘露醇10 g·L-1,Amp 50 mg·L-1;在对数生长期中期OD600为1.3时加入1 mmol·L-1 IPTG诱导,然后在30 ℃、180 r·min-1条件下继续培养9 h收获菌体;优化后目标蛋白CypA产量为66.7 mg·L-1发酵液,与优化前相比提高了76.6%.
Immobilization of Cells by Macro-porous NaCS-PDMDAAC Capsules and Cultivation in Shaking Flask and Bubble Bioreactor
大孔型NaCS-PDMDAAC微胶囊固定化细胞在摇床和鼓泡塔中的培养研究

ZHANG Jun,YAO Shan-Jing~,YING Xiao-Jiao,GUAN Yi-Xin,LIN Dong-Qiang,
张俊
,姚善泾,应小姣,关怡新,林东强

生物工程学报 , 2005,
Abstract: The membrane of sodium cellulose sulphate ( NaCS)-poly dimethyldiallylammonium chloride (PDMDAAC) microcapsule is compact and has low molecular weight cut-off, which would delay the mass transfer and affect the cell growth immobilized in the capsule. Macroporous NaCS-PDMDAAC microcapsules were prepared using the degradation of the starch by amylase in the membrane of the capsules. The pore size and the permeability in the membrane were improved obviously. As model cells, the Candida krusei CK1 and E. coli EC1 immobilized in the capsules were cultured in the shake flask and bubble column respectively. It was shown that the cell density immobilized in the microcapsules cultured in the bubble column was higher than that cultured in the shaking flask. It implied that the limiting factor of the cell growth in the capsule lied in the diffusion of the oxygen. Since the rate of the oxygen transporting across the membrane was greatly enhanced due to the enlarged pore size, the maximum cell density in the macroporous capsules was 20%-110% over than that in the standard capsules in the bubble column. However, the extent of E. coli cell density increasing was higher than that of the yeast, which may be due to the difference of the oxygen requirement between the two microbes.
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