An unconventional form of actin in protozoan hemoflagellate, Leishmania

Leishmania actin was cloned, overexpressed in baculovirus-insect cell system, and purified to homogeneity. The purified protein polymerized optimally in the presence of Mg2+ and ATP, but differed from conventional actins in its following properties: (i) it did not polymerize in the presence of Mg2+ alone, (ii) it polymerized in a restricted range of pH 7.0-8.5, (iii) its critical concentration for polymerization was found to be 3-4-fold lower than of muscle actin, (iv) it predominantly formed bundles rather than single filaments at pH 8.0, (v) it displayed considerably higher ATPase activity during polymerization, (vi) it did not inhibit DNase-I activity, and (vii) it did not bind the F-actin-binding toxin phalloidin or the actin polymerization disrupting agent Latrunculin B. Computational and molecular modeling studies revealed that the observed unconventional behavior of Leishmania actin is related to the diverged amino acid stretches in its sequence, which may lead to changes in the overall charge distribution on its solvent-exposed surface, ATP binding cleft, Mg2+ binding sites, and the hydrophobic loop that is involved in monomer-monomer interactions. Phylogenetically, it is related to ciliate actins, but to the best of our knowledge, no other actin with such unconventional properties has been reported to date. It is therefore suggested that actin in Leishmania may serve as a novel target for design of new antileishmanial drugs.     

Synthesis, biochemical evaluation of a range of potent 4-substituted phenyl alkyl imidazole-based inhibitors of the enzyme complex 17alpha-Hydroxylase/17,20-Lyase (P45017alpha)

We report the synthesis, biochemical evaluation and rationalisation of the inhibitory activity of a number of azole-based compounds as inhibitors of the two components of the cytochrome P-450 enzyme 17alpha-hydroxylase/17,20-lyase (P450(17alpha)), i.e. 17alpha-hydroxylase (17alpha-OHase) and 17,20-lyase (lyase). The results suggest that the compounds synthesised are potent inhibitors, with 7-phenyl heptyl imidazole (11) (IC(50)=320 nM against 17alpha-OHase and IC(50)=100 nM against lyase); 1-[7-(4-fluorophenyl) heptyl] imidazole (14) (IC(50)=170 nM against 17alpha-OHase and IC(50)=57 nM against lyase); 1-[5-(4-bromophenyl) pentyl] imidazole (19) (IC(50)=500 nM against 17alpha-OHase and IC(50)=58 nM against lyase) being the most potent inhibitors within the current study, in comparison to ketoconazole (KTZ) (IC(50)=3.76 microM against 17alpha-OHase and IC(50)=1.66 microM against lyase). Furthermore, consideration of the inhibitory activity against the two components shows that all of the compounds tested are less potent towards the 17alpha-OHase in comparison to the lyase component, a desirable property in the development of novel inhibitors of P450(17alpha). From the modelling of these compounds onto the novel substrate heme complex (SHC) for the overall enzyme complex, the length of the compound, along with its ability to undergo interaction with the active site corresponding to the C(3) area of the steroidal backbone, are suggested to play a key role in determining the overall inhibitory activity.     

磁选育浸矿菌种新方法的研究——磁泳分离菌种

世界无处不有磁,磁场对整个世界产生着重大的影响。本文通过大量镜检工作,观察到从酸性矿坑水中初步分离培养得到的部分细菌对外加磁场均有微弱的趋磁性。基于菌种的这种特性,设计了磁泳装置用不同的磁场梯度分离细菌,磁泳分离的方法可以初步分离出近磁、远磁茼,这两个菌群的生理特性有着很大的差异,主要体现在其对亚铁氧化和对金属高于的浸出上,远磁菌亚铁氧化活性比近磁菌高将近50％,远磁菌对铜离子的浸出效果也比近磁茼好。近磁菌在强磁性矿物培养基中生长情况较好,而远磁茼在弱磁性矿物培养基中生长情况较好。而且,在近磁茼的纯培养茼体中分离到磁性颗粒。实验结果证明。采用磁泳用于分离体内舍有磁性颗粒的细菌是可行并且有效的,这一分离技术和工艺的结合也将大大促进我国生物冶金的步伐。     

乳腺增生病动物模型的实验研究

目的探索建立与人类乳腺增生病相类似动物模型的最佳方法。方法采用手术、己烯雌酚、黄体酮处理动物。检测血液中雌二醇(E2)、黄体酮(P)、促卵泡生成激素(FSH)、促黄体生成激素(LH)水平;乳腺组织作组织形态学观察与评价。结果与对照组比较,各实验组动物的乳腺组织切片均有典型的乳腺增生;血清中各激素的水平也有不同的变化。结论采用“己烯雌酚 黄体酮”方法,以家兔为实验对象,建立乳腺增生病的动物模型是目前最好的方法。     

金钱鱼毒腺cDNA表达文库的构建及EST序列分析

以金钱鱼 (Scatophagusargus)毒腺为出发材料 ,构建以真核表达载体pcDNA3 0为基础的金钱鱼毒腺cDNA表达文库 .应用SMARTTM cDNALibraryConstruction技术和生物信息学分析等方法 ,通过对文库克隆的序列测定和初步生物信息学分析 ,获得了 2 0 1个金钱鱼新表达序列标签 (ESTs) ,其中已确定全长cDNA的克隆有 2 7个 ,包括多个核糖体大小亚基蛋白 (ribosomalprotein)、细胞凋亡相关蛋白 (programmedcelldeath 10 ) ,G蛋白信号调控子 (Gproteinsignalingregulator)、胸腺素(thymosinbeta 4 )、延伸因子 (translationelongationfactor 1 alpha)和泛素 (ubiquitin)等 .金钱鱼毒腺cDNA表达文库的成功构建 ,为研究金钱鱼毒腺的活性组分及其作用机制奠定了基础 ,也是分离新基因不可多得的重要资源     

草鱼胰岛素样生长因子-Ⅰ基因在大肠杆菌中的表达

 为构建草鱼 (Ctenopharyngodonidellus )胰岛素样生长因子 Ⅰ (IGF Ⅰ )大肠杆菌表达质粒 ,对已克隆到的草鱼IGF Ⅰ基因进行改造 .改造后的基因去除了原cDNA的信号肽和E区序列 ,并在基因的两端分别加入起始密码子和终止密码子 .将改造后的编码草鱼IGF Ⅰ成熟肽基因亚克隆到pBV 2 2 0中 ,构建成表达质粒pBVgIGF7.转化大肠杆菌 (Escherichiacoli)进行表达 .SDS PAGE显示 ,含重组表达质粒的菌株经热诱导后表达出一约 7 5kD的特异蛋白 .表达量占菌体总蛋白的2 0 0 3% ,表达产物主要以包涵体形式存在 .重组蛋白经纯化和复性后 ,采用MTT法测定其对草鱼吻端成纤维细胞PSF和草鱼卵巢细胞CO的促增殖作用 .结果表明 ,所获得的重组草鱼IGF Ⅰ具有生物活性     

细胞松弛素对嗜热四膜虫皮层细胞骨架蛋白的影响

测定了细胞松弛素B对嗜热四膜虫RF1株和BF5在生殖接合期的皮层骨架蛋白,尤其是在接合子接合膜形成和原核交换阶段影响甚大。作者发现VB处理后,与对照组相比较,皮层骨架蛋白146KD消失,27KD,43KD,47KD和174KD含量下降,32KD,41KD,51KD和54KD保持不变,结果显示,松弛素B对微纤毛蛋白27KD、43KD、47KD、146KD和174KD有影响。     

罗氏沼虾肌肉白浊病病原研究

从患白浊病的罗氏沼虾虾苗病灶分离到10多株菌株,其中一株经人工感染证实为病原菌,经ATB Expression自动细菌鉴定仪测试,为木糖葡萄球菌,其主要特性为拟球状（直径0.8-1.1μm）,革兰氏阳性,无鞭毛,无荚膜,不产生芽孢,发酵葡萄糖、果糖、麦芽糖、乳糖、海藻糖、阿拉伯糖、蔗糖及甘露醇产酸,β-半乳糖甘酶、过氧化氢酶及脲酶阳性,氧化酶,血浆凝固醇、V-P反应,精氨酸双水解酶、鸟氨酸脱羧酶均为阴性。最适生长温度、盐度和pH值范围分别为28-37℃、0%-2%、6-9,对红霉素、卡那毒素、头孢氨苄及庆大霉素等药物敏感。