Study on structure-activity relationship of mutation-dependent herbicide resistance acetohydroxyacid synthase through 3D-QSAR and mutation

Seventy-four sulfonylureas were synthesized and tested for their inhibitory activity against the whole enzyme of E. coli acetohydroxyacid synthase (AHAS, EC 2.2.1.6) isoenzyme II, and 3D-QSAR analyses were performed based on these inhibitory activities. The binding conformation of chlorimuron-ethyl, a commercial herbicide of AHAS, in the crystal structure of AHAS complex was extracted and used as template to build the initial three-dimensional structure of other sulfonylureas, and then all structures were fully geometry optimized. After systematic optimization of the alignment rule, molecular orienta- tion, grid space and attenuation factor, two satisfactory models with excellent performances (CoMFA: q2 = 0.735, r2 = 0.954, n = 7, r 2 pred = 0.832; CoMSIA: q2 = 0.721, r2 = 0.913, n = 8, r 2pred = 0.844) were estab- lished. By mapping the 3D contour maps of CoMFA and CoMSIA models into the possible inhibitory active site in the crystal structure of catalytic subunit of yeast AHAS, a plausible binding model for AHAS, with best fit QSAR in the literature so far, was proposed. Moreover, the results of 3D-QSAR were further utilized to interpret resistance of site-directed mutants. A relative activity index (RAI) for AHAS enzyme mutant was defined for the first time to relate the 3D-QSAR and resistance of mutants. This study, for the first time, demonstrated that combination of 3D-QSAR and enzyme mutation can be used to decipher the molecular basis of ligand-receptor interaction mechanism. This study refined our understanding of the ligand-receptor interaction and resistance mechanism in AHAS-sulfonylurea system, and provided basis for designing new potent herbicides to combat the herbicide resistance.     

Effects of niacin on nitric oxide synthase expression in rat lungs exposed to silica

The aim of this study was to evaluate the effects of niacin in diet on the expression of nitric oxide synthase (NOS) in rat lungs of the animal model of silicosis established by direct tracheal instillation of silica particles into rat lungs surgically. The niacin concentration in serum was analyzed by high performance liquid chromatography (HPLC). The expression of inducible nitric oxide synthase (iNOS) protein in paraffin-embedded lung sections was determined by streptavidin/peroxidase (SP) staining. Quantitative analysis by Image-Pro Plus was also performed on the expression of iNOS. The results showed that niacin concentration in serum of the niacin-treated rats was significantly higher than that in the control and silica-treated rats. After 7 days of silica instillation, iNOS integrated optical density (IOD) in rat lungs and total NOS and iNOS activities in bronchoalveolar lavage fluid (BALF) in silica-treated rats rose by 273420.75, 2.61 units/mL and 1.89 units/mL respectively, when compared with those in the control rats. Niacin treatment significantly reduced silica-induced iNOS IOD in rat lung tissues and total NOS and iNOS activities in BALF supernatant by 248292.35, 1.50 units/mL and 0.91 units/mL, respectively, as compared with those in silica-treated rats. Therefore, niacin can effectively attenuate the pathological expression of NOS in rat lung tissues induced by silica particles.     

运动对人体血清一氧化氮含量和一氧化氮合酶活性影响的研究

为了探讨人体在不同功能状态下一氧化氮（NO）和一氧化氮合酶（NOS）的变化，应用跑台提供运动负荷，分别在36名大学生进行了安静，亚极量负荷和力竭性运动后血清中NO和NOS的测定。结果发现，亚极量负荷下血清中NO含量显著高于安静状态时（P＜0．001），NOS活性极显著地低于安静状态时（P＞0．001），力竭性运动后，NO含量和NOS活性极显著低于亚极量负荷时（P＜0．001），并且NO已降至低于安静时的水平（P＞0．05），提示，人体血清中NOS活性随运动负荷的增大而持续下降；NO含量在运动的开始阶段随运动负荷的增大而升高，但当达到一定水平时，则随运动负荷的继续增大而下降。     

高原鼢鼠肌肉脂溶性物质对大鼠一氧化氮合成酶活性的影响

为了探讨高原鼢鼠肌肉脂溶性物质（FSC）的抗缺氧机制，40只SD大鼠随机分为4组：10％FSC组、10％FSC缺氧组、空白对照组和空白缺氧组。FSC组和空白组分别用10％FSC和蒸馏水连续灌胃30d，剂量为每天20mL/kg。灌胃结束后，10％FSC缺氧组和空白缺氧组大鼠在5000m海拔高度下，连续缺氧15d，每天8h，测定各组大鼠血清一氧化氮合成酶（NOS）活性。结果发现，在5000m海拔高度条件下，10％FSC在缺氧条件下能显著提高大鼠血清NOS活性和NO含量（P＜0.05)。     

查尔酮合酶基因的克隆及双元载体的构建

以中国水仙为材料，利用RT-PCR方法克隆查尔酮合酶基因(CHS)的cDNA，核酸序列分析表明，该基因的编码区长1167bp，编码389个氨基酸，通过进一步的中间克隆，将CHS连上CaMV35s启动子和Tnos终止子。进一步将重组片段插入植物表达载体pCAMBIA1301,PCR及测序结果都证明植物双元表达载体p1301BΩＣ构建获得成功。为进一步在植物中表达CHS奠定了基础。     

预防性使用HA对兔膝关节固定后关节软骨中iNOS表达的影响

为研究预防性使用透明质酸是否对兔膝关节固定后软骨中诱导性一氧化氮合酶(inducible nitric oxide synthase,iN-OS)的表达有下调作用,将24只雄性新西兰大白兔随机分成两组,每组12只.①透明质酸(hyaluronic acid,HA)组给予左膝关节内注射0.3 mLHA,每周1次,共行5次;②生理盐水(nature saline,NS)组给予左膝关节注射同等剂量NS,每周1次,共行5次.各组均于初次注射后将左下肢给予石膏管型固定,膝部预留窗口以备下次注射.于末次注射1周后处死动物,采集左膝关节股骨髁关节软骨制成石蜡切片.用免疫荧光方法标记各组软骨中的iNOS,在激光共聚焦显微镜下观察其表达情况,并用图像分析系统对其进行计数分析.结果HA组软骨iNOS表达的数量(22.617±3.263 5)明显低于NS组软骨(39.667±5.948 5).差别具有极显著统计学意义(P<0.01).说明:预防性使用HA对兔膝关节固定后软骨中iNOS的表达有下调作用.     

山新杨蔗糖合酶基因PCR介导的重组现象研究

[目的]山新杨(Poputlus davidiana ×P.bolleana)是林木基因工程育种基础和应用研究的良好材料;以山新杨蔗糖合酶基因(PdbSUS)为例研究聚合酶链式反应(polymerase chain reaction,PCR)过程介导的重组现象,在此基础上区分每一个PdbSUS基因的两种单倍型,为后续研...     

Mitochondrial association of alpha-synuclein causes oxidative stress

α-Synuclein is a neuron-specific protein that contributes to the pathology of Parkinson’s disease via mitochondria-related mechanisms. The present study investigated possible interaction of α-synuclein with mitochondria and consequences of such interaction. Using SHSY cells overexpressing α-synuclein A53T mutant or wild-type, as well as isolated rat brain mitochondria, the present study shows that α-synuclein localizes at the mitochondrial membrane. In both SHSY cells and isolated mitochondria, interaction of α-synuclein with mitochondria causes release of cytochrome c, increase of mitochondrial calcium and nitric oxide, and oxidative modification of mitochondrial components. These findings suggest a pivotal role for mitochondria in oxidative stress and apoptosis induced by α-synuclein. Received 27 December 2007; received after revision 7 February 2008; accepted 8 February 2008     

Effect of exogenous ammonium on glutamine synthetase, glutamate synthase, and glutamate dehydrogenase in the root of rice seedling

Root biomass of rice seedlings was increased at lower concentration of exogenous NH ₄ ⁺ , but it was decreased at higher concentration of exogenous NH ₄ ⁺ . The level of free NH ₄ ⁺ in the roots was accumulated gradually with the increase of NH ₄ ⁺ concentration in the nutrient solution. The content of the soluble proteins was essentially constant at higher NH ₄ ⁺ . The activities of glutamine synthetase (GS), NADH-dependent glutamate synthase (NADH-GOGAT), and NADH-dependent glutamate dehydrogenase (NADH-GDH) were risen with exogenous NH ₄ ⁺ concentration at the lower NH ₄ ⁺ concentration range. But the activities of GS and NADH-GOGAT were declined, and the level of NADH-GDH activity was kept constant under higher NH ₄ ⁺ concentration. The GS/GDH ratio suggested that NH ₄ ⁺ was assimilated by GS-GOGAT cycle under lower NH ₄ ⁺ concentration, but NADH-GDH was more important for NH ₄ ⁺ assimilation and detoxifying NH ₄ ⁺ to the tissue cells at the higher NH ₄ ⁺ level. According to the growth and the activity changes of these ammonium-assimilating enzymes of rice seedling roots, 10. 0 μg/mL NH ₄ ⁺ -N in nutrient solution was more suitable to the rice growth. Foundation item: Supported by the National Natural Science Foundation of China (No. 3987052), Natural Science Foundation of Hubei Province and International Rice Research Institute, P.O. Box. 3217, 1271 Makati City, Philippines. Biography: LI Ze-song(1968-), male, Graduate student.     

Microsomal glutathione S-transferases: selective up-regulation of leukotriene C4 synthase during lipopolysaccharide-induced pyresis

Cysteinyl-leukotrienes (cys-LTs) are potent smooth muscle contracting agents, which play key roles in inflammatory and allergic diseases. The committed step in cys-LT biosynthesis is catalyzed by leukotriene C₄ synthase (LTC4S) as well as microsomal glutathione S-transferase type 2 (MGST2) and type 3 (MGST3). Here we report that intraperitoneal injections of lipopolysaccharide in rats lead to a strong increase of LTC4S messenger RNA (mRNA) levels after approximately 1 h, particularly in the heart, brain, adrenal glands and liver, without any significant effect on MGST2 and MGST3 mRNA levels. After 6 h, LTC4S mRNA returns to basal levels, concomitant with a 4.9-, 4.0-, 2.9- and 2.3-fold induction of LTC4S protein in brain, heart, liver and adrenal gland, respectively. Hence, challenge with lipopolysaccharide in vivo causes an organ-selective, local priming for leukotriene C₄ synthesis. Moreover, these data suggest that LTC4S and cys-LTs may be involved in acute systemic inflammatory responses such as fever and tachycardia.Received 12 August 2004; received after revision 27 October 2004; accepted 1 November 2004