Salt-induced cooperativity in ATPase activity of plasma membrane-enriched fractions from cultured Citrus cells: kinetic evidence

ATPase activity was studied in plasma membrane-enriched fractions prepared from cultured Citrus sinensis L. cv. Osbeck cells. In general, properties of the plasma membrane ATPase from cultured cells, such as optimal pH and temperature. V_max and K_m were similar to those already observed in higher plants. The effects of high salt concentrations on ATPase activity were studied in membrane fractions derived from salt-sensitive and salt-tolerant cells grown in the presence or absence of salt. NaCl did not have an in vivo effect on V_max and the apparent K_m value for ATP. However, high concentrations of NaCl, or KCl, added in vitro, induced cooperativity in the enzyme and reduced the affinity of the enzyme for its substrate. Isoosmolar concentrations of sucrose or choline chloride failed to do so. Our results suggest that the plasma membrane ATPase of Citrus cells has more than one substrate-binding site on the native form of the enzyme which interact in the presence of salt and act independently in its absence.     

化学法合成生物素标记寡核苷酸探针

 生物素可通过氨烷基磷酰胺基团连接到寡核苷酸5'末端,反应在水溶液中进行,核苷酸的侧链基团不用保护。我们以这种化学法合成了生物素标记的寡核苷酸探针,其显色灵敏度达50pg,杂交灵敏度达0.4fmol,并与酶标生物素寡核苷酸探针进行了比较,也对两种不同显色体系作了比较。     

猴头菇对小鼠抗疲劳作用的实验研究

分别以猴头菇干粉（猴头菇Ⅰ组）和猴头菇浸出液（猴头菇Ⅱ组）饲喂小鼠,观察猴头菇对小鼠血清乳酸脱氢酶（ＬＤＨ）活力、血乳酸、血清尿素氮（ＢＵＮ）、肝糖原、肌糖原含量及运动耐力的影响。结果表明：实验６０ｄ后,猴头菇Ⅰ、Ⅱ组ＬＤＨ活力、肝糖原及肌糖原含量明显高于对照组（Ｐ＜０．０５或Ｐ＜０．０１）;运动后血乳酸的水平和ＢＵＮ的增量明显低于对照组（Ｐ＜０．０５或Ｐ＜０．０１）;运动后血乳酸消除速率显著高于对照组（Ｐ＜０．０５）;在运动耐力测定时在水中淹死的时间比对照组长得多（Ｐ＜０．０５）。提示：猴头菇具有明显的增强运动能力和解除疲劳的作用。     

大肠杆菌亮氨酰－tRNA合成酶LeuRS67R的纯化及其动力学研究

本实验室已得到的亮氨酰－ｔＲＮＡ合成酶（ＬｅｕＲＳ）基因,与文献相比,６７位氨基酸残基由Ｈｉｓ变为Ａｒｇ,此酶被定名为ＬｅｕＲＳ６７Ｒ。我们从该基因与ｐＵＣ１９重组质粒的大肠杆菌ＴＧ１转化子ＴＧ１－９１中得到ＬｅｕＲＳ的高表达,粗抽液中ＬｅｕＲＳ的表达量在转化子中比在宿主菌ＴＧ１中高２０倍以上。用三步拉层析得到电泳一条带的酶,其比活为１７８９单位／毫克。测定其动力学常数,氨酰化活力对Ｌｅｕ、ＡＴＰ的Ｋｍ值分别为０．０２７ｍｍｏｌ／Ｌ、０．４７ｍｍｏｌ／Ｌ,Ｋｃａｔ值分别为３．５～５．１ｓ－１。ＡＴＰ－ＰＰｉ交换活力对Ｌｅｕ、ＡＴＰ的Ｋｍ值分别为０．０３ｍｍｏｌ／Ｌ、１．０ｍｍｏｌ／Ｌ,Ｌｃａｔ值分别为１４０～１５５ｓ－１。此结果与从野生型大肠杆菌Ｋ－１２中提纯的ＬｅｕＲＳ的动力学常数差别很小,６７位氨基酸残基在与活性中心无直接关系的域可能是大肠杆菌的种间差异。     

肝癌组织中GD3合成酶的纯化

ＧＤ３合成酶是膜嵌合蛋白,定位于高尔基体,经过制作微粒体,Ｔｒｉｔｏｎ增溶、ＡＨ－Ｓｅｐｈａｒｏｓｅ及ＧＭ３－Ｇｌａｓｓｂｅａｄｓｅ及ＧＭ３－Ｇｌａｓｓｂｅａｄｓ亲和层析等步骤,二乙基亚硝胺诱发大鼠肝癌组织中的ＳＴ２被纯化了３１５９７倍,得率为０．３５％。ＳＤＳ－聚丙烯酰胺凝胶电泳后银染色呈１条蛋白着色带,分子量为５５ｋｄ。     

金花茶花药愈伤组织的体细胞减数分裂

金花茶(Camellia petelotii)的小孢子单核期花药,经培养和继代培养6个月后的愈伤组织中,发现有少量体细胞进行减数分裂。在减数第一和第二次分裂中,同源染色体的配对和分离基本正常,最后形成四分体。该愈伤组织经石蜡切片和压片观察,发现其主要由大量的液化细胞和贮藏细胞所组成,此外,还有少部分分生细胞。没有发现进一步的分化。其染色体数目,2n=30者占71.7%其余则为非整倍体。     

肝胆显像螯合物—氮基三乙酸单酰苯胺及其类似物的合成

本课题合成四个氮基三乙酸单酰苯胺(IDA)类及其类似物肝胆显像螯合物。(1)用家兔做了肝胆显像扫描实验,其显像效果较好。(2)～(3)为未见报道的化合物。     

高砷含水层参与腐殖酸-铁矿物转化的关键微生物群落及其对砷迁移转化的影响

【目的】探究不同腐殖酸浓度下参与含砷水铁矿转化的微生物类群组成和丰度变化及对砷释放的影响,预测原位高砷含水层中功能微生物群参与有机质—含砷铁矿物转化过程对砷转化释放的作用。【方法】对河套平原高砷地下水和同深度高砷沉积物中的铁还原功能群落进行富集培养,构建室内厌氧微宇宙体系,将富集菌群分别加入到实验室条件下合成的不同浓度腐殖酸(0、1.5、7、14 mg C/L)-含砷水铁矿体系中,通过体系中砷、铁形态及浓度的变化分析,结合高通量测序技术、X-射线衍射(X-ray diffractometer, XRD),探究不同条件下砷的释放固定和群落的演替。【结果】高砷地下水组(G组)和沉积物组(S组)富集得到的铁还原功能群落具有明显差异,G组中以Aeromonadaceae为特殊优势菌群,而S组中以Shewanellaceae为特殊优势菌群。微宇宙实验结果显示,S组的铁还原量相对较高且速率较快;G组与S组中液相砷形态存在明显差异,整个培养期内G组均以As(Ⅴ)为主,而S组中前期以As(Ⅴ)为主,当反应到达20 d时液相As(Ⅲ)高达3.4μmol/L,推测此时具有砷还原功能的群落占优势地位。当反应...     

A Rapid Determination of 30 Prohibited Pesticide Residues in Lycii Fructus by UPLC-MS/MS

Prohibited pesticide residues have become one of the main factors affecting the quality and safety of Lycii Fructus, However, rarely studies focus on the rapid determination of these residues. Here, a total of 30 kinds of prohibited pesticide residues were determined by ultra-performance liquid chromatography tandem triple quadrupole mass spectrometry (UPLC-MS/MS) in five different process ways. Pretreatment methods, chromatographic separation and detection conditions in mass spectrometry were all optimized accordingly. Among the five different pretreatment methods, the first and third solid phase extraction failed to provide high recoveries of sulfosulfuron compounds (both lower than 60%). Recovery of chlorphenamidine by the Quick Easy Cheap Effective Rugged and Safe multiresidue method (QuEChERS) was lower than 60%, which did not meet the requirements of trace determination. The concentrations of 30 prohibited pesticides residues treated by straightforward and solid phase extraction showed good linearity in their corresponding ranges, with correlation coefficients over 0.99. The average recoveries of straightforward ranged from 78.13% to 110.9%, while RSD ranged from 1.3% to 16.9%, albeit poor purification was observed. The recovery yield from solid phase extraction was between 67.75% and 103.08% with RSD value from 0.8% to 14.0%, which met the requirements of trace determination, this method has good precision and stability. These results could be employed to other Traditional Chinese Medicines (TCMs) in detecting prohibited pesticide residues.