宏基因组来源耐Mn2+、热稳定细菌漆酶的分子克隆及酶学特性

【背景】漆酶和锰过氧化物酶(Manganese peroxidase,Mnp)是木质素降解的主要酶,二者有协同效应。Mnp活性依赖于Mn~(2+),而Mn~(2+)是大多数漆酶的抑制剂。【目的】获得耐Mn~(2+)的细菌漆酶用于木质素降解。【方法】构建许昌市某污水河污泥宏基因组文库,通过活性筛选获得细菌漆酶基因lac1542。使用大肠杆菌异源表达Lac1542,研究纯化后的重组蛋白酶学性质并进一步检测了含Lac1542复合酶系降解木质素能力。【结果】测序结果显示lac1542编码一个含513个氨基酸的蛋白。以ABTS为底物Lac1542最适反应pH为4.0,在pH 3.0-6.5范围内酶活性稳定。最适反应温度是75°C,在70°C以下酶活性稳定;100 mmol/L的Mn~(2+)仍能提高酶的活性。动力学参数研究发现,该酶的最适底物顺序为:ABTS丁香醛联氮儿茶酚2,6-DMP愈创木酚。Lac1542/Mnp复合酶系对木质素降解率为47.8%,比单独使用Mnp木质素降解率(22.4%)提高25.4%。Lac1542/Mnp/灰盖鬼伞过氧化物酶(Coprinus cinereus Peroxidase,CIP)复合酶系木质素降解高达71.5%,比Mnp/CIP酶系木质素降解率(48.9%)提高22.6%,加入Lac1542后的复合酶系能明显提高木质素的降解率。【结论】Lac1542的可溶性表达、耐受高浓度Mn~(2+)、热稳定性使得Lac1542可以替代一些经典的真菌漆酶应用于制浆、造纸、纤维素乙醇生产、染料脱色等工业。     

微藻光密度与细胞密度及生物质的关系

以四种常见微藻,小球藻(Chlorella sp.XQ-20044)、栅藻(Scenedesmus sp.SS-200716)、绿球藻(Chlorococcum sp.)和螺旋藻(Spirulina sp.CH-164)为实验材料,用梯度稀释法测定对数生长期不同浓度藻液的光密度(OD)、细胞密度和生物质干重(DW),在光自养分批培养模式下对4种微藻进行OD-波长(350—800 nm)扫描,同时测定细胞密度和生物质干重,分析藻液OD与细胞密度、生物质干重的关系。结果表明:在任何波长下,对数生长期的4种微藻细胞密度与OD值、生物质干重与OD值的变化都不成比例,波长不同其拟合曲线偏离直线的程度不同。但是,在435 nm处这种关系最接近直线,可以用直线方程近似描述(R20.98),其它波长处细胞密度-OD、干重-OD的关系都可以用二项式方程很好地描述(R20.99)。因此,光密度法适用于连续和半连续培养,可以用435 nm处测得的OD值计算细胞密度与干重。但是在分批培养模式下,4种微藻DW/OD比值随着培养时间均逐渐上升。小球藻DW/OD540为0.19—0.44 g/L,栅藻DW/OD540为0.36—0.53 g/L,绿球藻DW/OD540为0.48—0.75 g/L,螺旋藻DW/OD560为0.46—0.74 g/L,因此分批培养模式下采用测定藻液OD值反映细胞密度和生物质的方法不适用,只有直接测定细胞密度和生物质才是准确的。研究结果为正确使用分光光度法监测微藻生长提供依据。     

Embryology of Plagiopteron suaveolens Griffith (Plagiopteraceae) and its systematic implications

A study of the embryology of Plagiopteron suaveolens Griffith is presented in order to evaluate the affinities of the genus. Anther wall formation is of the dicotyledonous type. The anther tapetum is glandular and its cells become two to four nucleate. Cytokinesis in the microspore mother cell is simultaneous and the resultant tetrad is tetrahedral. The mature pollen grains are two-celled. The anther has 4-locules at anthesis. The ovule is anatropous, bitegmic and weakly crassinucellate with only one cell-layered parietal tissue. The micropyle is zig-zag in section and formed by both inner and outer integuments which are three and three to four cell layers thick respectively. The megaspore archesporium is one-celled and the embryo sac development is of the Polygonum type. An endothelium is formed with early disintegration of nucellar tissue. The antipodals multiply to 15 to 20 cells. Embryological evidence supports the placement of Plagiopteron in its own family, Plagiopteraceae, with closest relationship with Celastraceae and close relationship with Elaeocarpaceae, Rhizophoraceae and some families of the Geraniales.     

Malaria transmission blocking immunity and sexual stage vaccines for interrupting malaria transmission in Latin America

Malaria is a vector-borne disease that is considered to be one of the most serious public health problems due to its high global mortality and morbidity rates. Although multiple strategies for controlling malaria have been used, many have had limited impact due to the appearance and rapid dissemination of mosquito resistance to insecticides, parasite resistance to multiple antimalarial drug, and the lack of sustainability. Individuals in endemic areas that have been permanently exposed to the parasite develop specific immune responses capable of diminishing parasite burden and the clinical manifestations of the disease, including blocking of parasite transmission to the mosquito vector. This is referred to as transmission blocking (TB) immunity (TBI) and is mediated by specific antibodies and other factors ingested during the blood meal that inhibit parasite development in the mosquito. These antibodies recognize proteins expressed on either gametocytes or parasite stages that develop in the mosquito midgut and are considered to be potential malaria vaccine candidates. Although these candidates, collectively called TB vaccines (TBV), would not directly stop malaria from infecting individuals, but would stop transmission from infected person to non-infected person. Here, we review the progress that has been achieved in TBI studies and the development of TBV and we highlight their potential usefulness in areas of low endemicity such as Latin America.     

能在肝癌细胞特异表达的自主复制型肝导向基因转移系统

构建了以人甲胎蛋白基因上游调控序列控制外源基因的表达的自主复制型EB病毒复制子载体pEBAF.该载体与半乳糖基化组蛋白结合组成了一种能为肝细胞表面特异受体识别并内在化的核酸蛋白复合物,能通过肝细胞受体介导的内吞作用以非病毒感染的形式,将外源基因导入细胞内自主复制并仅在产甲胎蛋白的肝癌细胞特异表达.这种新型的基因转移系统,具有比直接使用重组病毒颗粒感染细胞更高的安全性,有应用于原发性肝癌的基因治疗的前景     

Respiration during recovery from exercise: effects of trapping and release of femoral blood flow

To investigate the contribution of vascular and metabolic stimuli to the sustained hyperpnea after exercise, the respiratory effects of obstructing and then releasing the femoral blood flow were recorded in 15 normal volunteers during recovery from steady-state cycle exercise (80 W). Obstruction was achieved using cuffs around the upper thighs, inflated for the first 2 min of recovery to a pressure of 200 mmHg. Cuff inflation significantly reduced ventilation during recovery compared with control (P less than 0.001); the subsequent release of pressure was accompanied by an increase in ventilation (averaging 3.2 l/min), which began on the first breath after release. This preceded a rise in end-tidal CO2 (maximum 8.3 Torr increase), which first became significant on the fourth breath after release and led to a further rise in ventilation. The first-breath increase in ventilation after cuff release persisted, although slightly attenuated (averaging 2.5 l/min), in additional experiments with inspired O2 fraction of 1.0. The pattern of ventilatory response was also similar when the experiments were performed with 5% CO2 in air as the inspirate. The immediate rise in ventilation on cuff release, together with the persistent response on 100% O2, suggests that the vascular changes resulting from cuff release exert an influence on ventilation independent of the effects of released metabolites on the known chemoreceptors. The persistence of the response on 5% CO2 indicates that CO2-sensitive lung afferents do not have a major role in these responses.     

丙型肝炎病毒核心蛋白抑制乙型肝炎病毒转录的分子机理

为研究丙型肝炎病毒 (HCV)核心蛋白抑制乙型肝炎病毒 (HBV)表达的分子机理 ,从HCV和HBV共感染中国病人血清中分离了 5个含HCV 5′非编码区 ( 5′NCR)和核心区 (CR)cDNA序列的克隆 .序列比较和系统发育分析显示 :从共感染病例中分离的HCV序列与其它已发表的从单独HCV感染病例中分离的序列没有显著差异 .共转染实验证实采用从其中 1例共感染病人中分离的核心蛋白cDNA基因所表达的核心蛋白 ,可抑制HBV表面抗原 (HBsAg)和HBVe抗原 (HBeAg)的表达 ,缺失突变分析说明HCV核心蛋白的C端疏水区对这种抑制作用是必需的 ,报道基因产物分析进一步说明了HBVC启动子和增强子Ⅱ是HCV核心蛋白的效应靶序列之一 ,而HCV核心蛋白在肝细胞和非肝细胞中均对HBV和其他细胞和病毒的基因的转录起负调节作用 .因此 ,认为HCV核心蛋白是一种多功能的负调节因子 ,与HCV和HBV以及HCV与细胞之间的相互作用密切相关     

一个含端粒序列的水稻BAC克隆的分析和定位

对一个含 (TTTAGGG) _n 同源序列的水稻BAC克隆 (BAC2 )进行了分析 ,揭示出水稻近端区DNA的组成 .BAC2的插入片段中除含有大量的以串联形式存在的称之为TA35 2序列的卫星DNA外 ,还含有TTTAGGG或其变体组成的简单重复 .荧光原位杂交 (FISH)将含 (TTTAGGG) _n 序列的 0 .8kbPstⅠ片段定位在至少 5对染色体的端粒区 .通过对BAC2中低拷贝序列的RFLP分析 ,BAC2被定位在水稻第 6号染色体端部 .这些结果说明水稻的端粒序列可能也是TTTAGGG或其变体构成的简单重复 ,而与其相连的卫星DNATA35 2则属于端粒相关序列 .