抗厌氧菌中草药的系列研究：Ⅱ.桂皮醛抗厌氧菌的实验研究

为探索肉桂在防治厌氧菌感染应用于临床的可能性,作者进一步用122株厌氧菌检测肉桂的抗菌活性。肉桂起抗菌作用的主要成分是桂皮醛。本文用商品桂皮醛,加吐温80助溶,配成含桂皮醛512μg/ml、256μg/ml 等一系列培养液,进行试管法测定 MIC、MBC。结果 MIC 在128μg/ml 以下者占所试菌株的76%,在256μg/ml 以下占96.7%。脆弱类杆菌、产黑素类杆菌相对更敏感。MBC 一般显著高于 MIC,因此桂皮醛主要是抑菌作用。从细菌的形态学观察,推论桂皮醛主要是作用细菌的胞壁,鉴于桂皮醛有一定毒性,作者建议可作为局部抗厌氧菌药物。     

Na~(+)、Ca~(2+)、Cu~(2+)和Zn~(2+)与HSA相互作用的~(23)(Na)-NMR研究

 本文应用~23Na-NMR波谱技术,研究了Na~(+)、Ca~(2+)、Cu~(2+)和Zn~(2+)与人体血清白蛋白(HSA)的相互作用。在实验基础上,通过引入两位快交换模型,拟合计算获得了Na~(+)与HSA相互作用的结合常数和处于结合状态Na~(+)的相关时间;实验表明Ca~(2+)能与Na~(+)竞争同HSA结合,拟合计算获得了两者与HSA相互作用结合常数的比值,棕榈酸钠能增强Ca~(2+)同Na~(+)竞争与HSA结合的能力;从实验上未能观察到Cu~(2+)、Zn~(2+)能同Na~(+)竞争与HSA相互作用的证据。     

冬虫夏草多糖的分子结构与免疫活性

 冬虫夏草多糖(在本文中名为CS-81002)是由冬虫夏草菌Cordyceps sinensis(Berk)Sacc在人工发酵条件下产生的胞外多糖。用凝胶过滤法测出CS-81002的分子量Mr=43kD。CS-81002的单糖组成为Man:Gal:Glc=10.3:3.6:1。甲基化分析和部份酸水解结果表明,CS-81002是多分支的杂多糖,由→6)-Manp-(1→形成主链,大约每10个主链Man残基中,有6个在C3位上被取代(即形成→3,6)-Manp-(1→分支),有4个在位C2上被取代(即形成→2,6-Manp-(1→分支),从而形成侧链。主链中还有少许未被取代的Man残基。侧链则由→3-)-Galf-(1→,→4)-Glcf-(1→,→4)-Manp-(1→和→2)-Manp-(1→组成。位于非还原末端的,则三种单糖残基都有。CS-81002在5mg/kg体重×12剂量条件下,对正常的昆明小鼠腹腔巨噬细胞吞噬功能有显著促进作用。在同样剂量条件下,对正常LACA小鼠脾脏溶血斑形成细胞(PFC)对绵羊红细胞的溶血作用无显著影响。不同程度的部份酸水解可使CS-81002的分子量下降,分支减少,并且对巨噬细胞吞噬功能的促进作用亦有下降趋势。在所得到的各个部份酸水解级分(分子量分别为41000,40000,32000,16000和12000)中,分子量为12000的级分对巨噬细胞吞噬功能无促进作用。     

中国鲎凝集素的分离纯化及性质研究

 经N-乙酰氨基葡萄糖交联琼脂糖亲和层析及以交联琼脂糖介质的高效液相分子筛层析,从中国鲎细胞溶解物中分离纯化了一种凝集素,其活性比原料鲎试剂提高128倍。鲎凝集素SDS电泳时表现出分子量为69000,和72000的二个亚基。N-乙酰氨基葡萄糖、D-半乳糖,D-甘露糖及岩藻糖等对鲎凝集素凝集鸡红细胞的活性有显著抑制作用,加热60℃,10分钟可使凝集素活性基本丧失。CaCl_2为凝集素活性所必需。鲎凝集素与肺炎球菌C多糖有沉淀反应。     

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。     

外源脂肪酸及胆固醇对莱氏衣原体膜糖脂含量的影响

 <正> 单葡萄糖甘油二酯(MGDG)和双葡萄糖甘油二酯(DGDG)是莱氏衣原体膜上主要的极性脂。糖脂的生理功能过去很少研究,至今仍不清楚。近年来实验结果表明,MGDG在膜上容易形成六角形Ⅱ结构,而DGDG则形成脂双层结构。六角形Ⅱ结构的出现与生物膜的生理功能的关系是当前瞩目的研究内容。本文首次研究了外源脂肪酸和胆固酵对莱氏衣原体AIH089菌株膜上糖脂含量的影响。     

Expression of the human retinoblastoma gene product pp110RB in insect cells using the baculovirus system

The product of the retinoblastoma susceptibility gene (RB) was overproduced in cultured insect cells using the baculovirus expression system. Upon insertion of the cloned human RB complementary DNA sequence into the viral genome downstream of the promoter of the polyhedrin gene, full-length RB protein with an apparent molecular weight of 110,000 was expressed in the insect cells. This protein was found to be phosphorylated, located in the nuclei of the infected cells, and immunologically indistinguishable from pp110RB of human cells as assayed by several anti-RB antibodies. Following cell disruption and a one-step immunoaffinity chromatographic purification, 6-12 mg of soluble pp110RB with approximately 95% purity were obtained per liter of infected suspension culture. Characterization of the two known biochemical properties of RB protein showed that this purified protein from insect cells behaved similarly to the authentic human pp110RB. First, it bound to DNA, and second, it could form a specific complex with SV40 T antigen in vitro. Prompt translocation of the protein from cytoplasm to nucleus after microinjection further indicated that the purified RB protein may be active. The availability of soluble, intact, and presumably active pp110RB in large quantity represents a significant advance for studying the biochemical and biophysical properties of the RB gene product as well as its potential biological function in cancer suppression.     

Prolonged gene expression and cell survival after infection by a herpes simplex virus mutant defective in the immediate-early genes encoding ICP4, ICP27, and ICP22.

Very early in infection, herpes simplex virus (HSV) expresses four immediate-early (IE) regulatory proteins, ICP4, ICP0, ICP22, and ICP27. The systematic inactivation of sets of the IE proteins in cis, and the subsequent phenotypic analysis of the resulting mutants, should provide insights into how these proteins function in the HSV life cycle and also into the specific macromolecular events that are altered or perturbed in cells infected with virus strains blocked very early in infection. This approach may also provide a rational basis to assess the efficacy and safety of HSV mutants for use in gene transfer experiments. In this study, we generated and examined the phenotype of an HSV mutant simultaneously mutated in the ICP4, ICP27, and ICP22 genes of HSV. Unlike mutants deficient in ICP4 (d120), ICP4 and ICP27 (d92), and ICP4 and ICP22 (d96), mutants defective in ICP4, ICP27, and ICP22 (d95) were visually much less toxic to Vero and human embryonic lung cells. Cells infected with d95 at a multiplicity of infection of 10 PFU per cell retained a relatively normal morphology and expressed genes from the viral and cellular genomes for at least 3 days postinfection. The other mutant backgrounds were too toxic to allow examination of gene expression past 1 day postinfection. However, when cell survival was measured by the capacity of the infected cells to form colonies, d95 inhibited colony formation similarly to d92. This apparent paradox was reconciled by the observation that host cell DNA synthesis was inhibited in cells infected with d120, d92, d96, and d95. In addition, all of the mutants exhibited pronounced and distinctive alterations in nuclear morphology, as determined by electron microscopy. The appearance of d95-infected cells deviated from that of uninfected cells in that large circular structures formed in the nucleus. d95-infected cells abundantly expressed ICP0, which accumulated in fine punctate structures in the nucleus at early times postinfection and coalesced or grew to the large circular objects that were revealed by electron microscopy. Therefore, while the abundant accumulation of ICPO in the absence of ICP4, ICP22, and ICP27 may allow for prolonged gene expression, cell survival is impaired, in part, as a result of the inhibition of cellular DNA synthesis.     

Analysis of the serotype-specific epitopes of avian infectious bronchitis virus strains Ark99 and Mass41.

The Ark and Mass serotype-specific epitopes of infectious bronchitis virus were studied by immunofluorescence and immunoprecipitation of mutant and recombinant spike glycoproteins (S protein) expressed in mouse L cells. Serotype-specific monoclonal antibodies could bind to the recombinant proteins of Ark99 and Mass41 expressed from the chimeras in which the N-terminal thirds of the S1 sequences were reciprocally exchanged. Therefore, it appears that the respective serotype-specific epitopes of both strains were localized within the C-terminal two-thirds of the S1 proteins. Deletion and insertion of a five-amino-acid fragment on the S1 proteins of Ark99 and Mass41, altered the serotype-specific epitopes. This result implies that the five-amino-acid insertion on the S1 protein of the Ark serotype was involved in determining the conformation of the protein, probably acting as a spacer. In addition, it appears that an interaction between sequences of the N-terminal third and the remaining portion of the S1 protein determines the tertiary structure of the protein as well as the conformation of the serotype-specific epitope.