细菌肽聚糖、磷壁酸的教学探讨

由于构成肽聚糖四肽链的氨基酸组成不同,交联方式又变化多端,从而不同种类细菌的肽聚糖结构多种多样。在描绘肽聚糖结构时要注意:1.四肽链第二位的D-谷氨酸与下一个氨基酸的氨基连接成肽键的是r-羧基,而不是α-羧基。2.交联主要是两条邻近的不同聚糖链骨架上与N-乙酰胞壁酸相连的两条相邻四肽链间的交联。3.G-细菌是两条四肽链直接交联,没有桥肽的参与。溶菌酶仅水解N-乙酰胞壁酸c-1和N-乙酰葡糖胺c-4之间的β-1,4糖苷键;而青霉素是抑制肽聚糖合成的最后阶段——转肽反应。由于两者的作用机理完全不     

人源溶菌酶结构与功能的研究进展

人溶菌酶是一类人体内天然存在的能够溶解细菌细胞壁的碱性蛋白的总称。其作用特征是能够裂解肽聚糖中的N-乙酰氨基葡萄糖与N-乙酰氨基甲酸之间的β-(1,4)-糖苷键。人溶菌酶具有抗菌、抗炎、抗病毒和增强免疫力等多种特性,因此在国内外市场上应用广泛。本文就人溶菌酶的结构特点、表达部位、功能表达以及应用情况进行综述。     

肾综合征出血热纯化疫苗的SDS-PAGE分析

为了证明蛑综合征出血热纯化疫苗的主要成分坦病毒蛋白,采用出血热纯化疫苗经浓缩后进行SDS－PAGE和Western-blotting分析。结果 经SDS－PAGE显示,肾综合征出血热纯化疫苗有三条蛋白带,分子量分别约为70kD、55kD和50kD,与汉坦病毒三种结构蛋白（糖蛋白G1、G2和核蛋白NP）的分子量相符;经Western-blotting显示,分子量50kD的蛋白带反应阳性,分子量70kD和55kD的蛋白带无反应,认定出血热纯化疫苗的主要成分为汉坦病毒蛋白,主要由G1、G2和NP三种结构蛋白构成。     

蜀柏毒蛾核型多角体病毒结构多肽及基因组酶切分析

对蜀柏毒蛾核型多角体病毒(Parocneria orienta Nuclear polyhedrovirus,简称PaorNPV)形态结构、结构多肽、限制性内切酶图谱等特性进行了研究.采用不连续系统垂直板SDS-PAGE分析了PaorNPV的多角体蛋白、病毒粒子结构多肽.应用5种限制性内切酶对PaorNPV基因组DNA进行了酶切分析.结果表明:经热处理的多角体蛋白仅有一条带,分子量为31.5 kD,不经热处理的多角体蛋白有三条带,分子量分别为31.5 kD、29.1 kD、28.6 kD;病毒粒子包含有25种结构多肽,分子量范围在17.6-114.6 kD之间.PaorNPV DNA经BamH I.EcoR I、HindⅢ、Pst I和Xho I酶切分别产生9、12、12、12和14条片段.基因组大小平均为124.6 kb.     

大肠杆菌细胞壁肽聚糖的化学修饰及荧光标记

【目的】发展一种活细菌细胞壁荧光标记方法,为后期研究细菌肽聚糖的生物合成和代谢规律以及其与细菌感染致病的关系提供新的工具。【方法】对细菌肽聚糖的生物合成前体N-乙酰葡萄糖胺-1-磷酸(GlcNAc-1-P)进行化学修饰,设计并合成含有叠氮基的GlcNAc-1-P类似物(化合物5:Ac3GlcNAz-1-P)。将该类似物与细菌共同孵育,使其作为探针进入细菌肽聚糖天然合成途径。之后提取并酶解肽聚糖组分,用红外光谱(FTIR)和液质联用(LC/MS)检测探针是否通过代谢进入细菌肽聚糖结构中。同时用外源荧光素对代谢掺入细菌肽聚糖中的探针进行染色。在激光共聚焦显微镜下观察对活细菌的荧光标记效果。【结果】通过四步有机合成反应,以79%的总收率成功获得了化合物5。将大肠杆菌(Escherichia coli BL21)作为模式菌株与化合物5共孵育后,其肽聚糖组分的LC/MS和FTIR分析结果均显示探针可以被细菌利用并被代谢掺入到肽聚糖结构中。激光共聚焦显微镜观察结果显示,荧光素可以高效标记表面携带有生物正交探针的大肠杆菌。【结论】设计合成了一种新型探针,可用于活细菌成像,为深入研究细菌肽聚糖的生物学功能及其与细菌感染致病的关系提供了一种简便的方法。     

汉滩病毒核蛋白的分段表达及抗原表位分析

在大肠杆菌中对汉滩病毒S基因4种不同长度片段的重组表达质粒进行诱导表达.结果表明表达的4种GST-NP融合蛋白均以不溶性包含体形式存在于菌体细胞内,表达量分别占菌体蛋白总量的29-36%,分子量分别约为72 kD、66 kD、54 kD和44 kD.Western blot显示54 kD和72 kD融合蛋白用酶标抗汉滩病毒NP McAb 1A8和抗GST McAb 3C11染色呈阳性反应.66 kD和44 kD融合蛋白仅与酶标3C11呈阳性反应.用凝血酶切去GST,获得4种汉滩病毒重组核蛋白(rNP),分子量分别约为44 kD、40 kD、26 kD和16 kD.用19株McAb对表达产物做抗原位点分析,结果完整的rNP可与19株McAb中的13株反应,McAb反应谱与天然NP相同;S1.1 kb表达产物(N-端1-37和C端402-429位aa缺失)和S 0.5 kb表达产物(N-端1-274位aa缺失)与19株McAb均不发生反应;S0.7 kb表达产物(C-端275-429位aa缺失)可与5株组特异性McAb反应.表明汉坦病毒核蛋白上的抗原位点主要存在于N-端1-37位aa区段内.     

斑玉蕈Hypsizygus marmoreus凝集素的部分性质和细胞凝集活性分析

斑玉蕈子实体经生理盐水抽提、30%~60%饱和度的硫酸铵沉淀、DEAE-Cellulose和SephadexG-100柱层析纯化得到斑玉蕈凝集素(HypsizygusmarmoreusLectin,简称HML)。HML经PAGE显示单一条带,SDS-PAGE测得亚基相对分子质量为34.2kD,SephadexG-100凝胶过滤测得相对分子质量为35kD,中性糖含量为7.2%,含有17种氨基酸,IEF-PAGE测得其等电点为8.15。该凝集素能凝集多种动物红细胞和人的A、B、AB和O血型红细胞,对兔红细胞的凝集作用可被甘露糖、半乳糖、N-乙酰半乳糖胺和岩藻糖所抑制。HML对热较敏感,经50℃处理10min,凝集活性下降明显,但对酸碱具有一定的稳定性,在pH5.0~8.0范围内较稳定。Β-消去反应测得其糖和蛋白质的连接键为O-型糖肽键。     

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。     

斑玉蕈Hypsizygus marmoreus凝集素的部分性质和细胞凝集活性分析

斑玉蕈子实体经生理盐水抽提、30%~60%饱和度的硫酸铵沉淀、DEAE-Cellulose和SephadexG-100柱层析纯化得到斑玉蕈凝集素(HypsizygusmarmoreusLectin,简称HML)。HML经PAGE显示单一条带,SDS-PAGE测得亚基相对分子质量为34.2kD,SephadexG-100凝胶过滤测得相对分子质量为35kD,中性糖含量为7.2%,含有17种氨基酸,IEF-PAGE测得其等电点为8.15。该凝集素能凝集多种动物红细胞和人的A、B、AB和O血型红细胞,对兔红细胞的凝集作用可被甘露糖、半乳糖、N-乙酰半乳糖胺和岩藻糖所抑制。HML对热较敏感,经50℃处理10min,凝集活性下降明显,但对酸碱具有一定的稳定性,在pH5.0~8.0范围内较稳定。β-消去反应测得其糖和蛋白质的连接键为O-型糖肽键。     

意大利蜜蜂N-乙酰-β-D-氨基葡萄糖苷酶的分离纯化及性质分析

【目标】N-乙酰-β-D-氨基葡萄糖糖苷酶(NAGase)是一种重要的几丁质分解酶,能从N-乙酰葡萄糖苷的非还原端催化去除β-1,4-N-乙酰-D-氨基葡萄糖残基,参与了昆虫外骨骼的蜕皮过程。研究蜜蜂该酶的特征有助于阐明其在蜜蜂发育过程中的作用机制。【方法】采用40%-70%硫酸铵分级沉淀、DEAE-纤维素离子交换层析和葡聚糖G-100凝胶过滤层析的方法从意大利蜜蜂Apis mellifera ligustica幼虫体内分离纯化NAGase。以对-硝基苯-N-乙酰-β-D-氨基葡萄糖苷(pNP-NAG)为底物检测该酶的活力,用native PAGE和SDS-PAGE检测酶的纯度。IEF-PAGE测定该酶等电点。葡聚糖G-200凝胶过滤层析测定酶的总分子量。【结果】结果显示,纯化的NAGase酶的比活力为803. 09 U/mg,总分子量为77. 3 kD。结合SDS-PAGE表明该酶由两个具有相同分子量(39 k D)的亚基组成。该酶等电点为4. 8。酶水解底物pNP-NAG的过程遵循米氏方程,米氏常数(Km)和最大反应速度(Vm)分别为0. 11 mmol/L和17. 65μmol/L·min。该酶水解反应的最适pH和最适温度分别为pH 5. 5和60℃。酶催化pNP-NAG反应的活化能为64. 8 k J/mol。Pb2+,Cu2+,Zn2+和Al3+对该酶有不同程度的抑制作用。【结论】本研究描述了意大利蜜蜂NAGase的分离纯化方法及其理化性质,为进一步进行蜜蜂NAGase的结构解析和功能研究奠定基础。     

Recent advances in the study of Kaposi's sarcoma-associated herpesvirus replication and pathogenesis

It has now been over twenty years since a novel herpesviral genome was identified in Kaposi's sarcoma biopsies. Since then, the cumulative research effort by molecular biologists, virologists, clinicians, and epidemiologists alike has led to the extensive characterization of this tumor virus, Kaposi's sarcoma-associated herpesvirus(KSHV; also known as human herpesvirus 8(HHV-8)), and its associated diseases. Here we review the current knowledge of KSHV biology and pathogenesis, with a particular emphasis on new and exciting advances in the field of epigenetics. We also discuss the development and practicality of various cell culture and animal model systems to study KSHV replication and pathogenesis.     

The structure, reproduction and taxonomy of Vidalia and Osmundaria (Rhodophyta, Rhodomelaceae)

Comprises species occurring mostly in subtidal habitats in tropical, subtropical and warm-temperate areas of the world. An analysis of the type species, V. spiralis (Sonder) Lamouroux ex J. Agardh, a species from Australia, establishes basic characters for distinguishing species in the genus. These characters are (1) branching patterns of thalli, (2) flat blades that may be spiralled on their axis, (3) width of the blade, (4) primary or secondary derivation of sterile and fertile branchlets and (5) position of sterile and fertile branchlets on the thalli. Application of the latter two characters provides an important basic method for separation of species into three major groups. Osmundaria , a genus known only in southern Australia, was studied in relation to Vidalia , and its separation from the Vidalia assemblage is not accepted. Species of Vidalia therefore are transferred to the older genus name, Osmundaria. Two new species, Osmundaria papenfussii and Osmundaria oliveae are described from Natal. Confusion in the usage of the epithet, Vidalia fimbriala Brown ex Turner has been clarified, and Vidalia gregaria Falkenberg, described as an epiphyte on Osmundaria pro/ifera Lamouroux, is revealed to be young branches of the host, Osmundaria prolifera.     

New or rare chromosome counts in Artemisia L. (Asteraceae, Anthemideae) and related genera from Kazakhstan

Fifteen chromosome counts of six Artemisia taxa and one species of each of the genera Brachanthemum, Hippolytia, Kaschgaria, Lepidolopsis and Turaniphytum are reported from Kazakhstan. Three of them are new reports, two are not consistent with previous counts and the remainder are confirmations of very scarce (one to four) earlier records. All the populations studied have the same basic chromosome number, x = 9, with ploidy levels ranging from 2x to 6x. Some correlations between ploidy level, morphological characters and distribution are noted.     

肝癌中HBV和HCV基因和抗原的分布及意义

采用原位分子杂交方法检测HCV RNA及HBV X基因;采用免疫组织化学方法研究HCV核心抗原,非结构区C33c抗原及HBxAg在肝细胞肝癌中的定位及分布.结果表明(1)HCV RNA、HBV X基因在肝细胞肝癌组织检出率分别为40%(55/136)和82%(112/136).HCV RNA定位于癌细胞的胞浆内,阳性细胞呈散在、灶状及弥漫分布三种形式;HBV X基因在肝癌细胞中的分布呈胞浆型、核型及核浆型,阳性细胞也呈上述三种分布形式;(2)HCV C33c抗原、核心抗原在肝细胞肝癌中的阳性率为81%(133/164)及86%(141/164).C33c抗原定位于癌细胞及肝细胞的胞浆内;核心抗原既定位于癌细胞核中,又可定位于胞浆中.C33c抗原阳性细胞以灶状分布为主;而核心抗原阳性细     

Selection with two alleles and complete dominance

For a plant selection model with frequency-independent viabilities, fertilities and selfing rates, it is shown that apart from global fixation, for certain parameter combinations a protected polymorphism and facultative fixation (either allele may become fixed according to initial frequencies) may both occur. Facultative fixation requires different selling rates for the dominant and recessive type. Protection of the polymorphism requires resource allocation for male and female function. In this connection the problem of purely genetically caused population extinction is discussed.
For general frequency dependence and regular segregation, the chances for establishment of a completely recessive gene are compared to those of a completely dominant gene. It is proven that the process of establishment of the recessive gene, despite a fitness advantage, may be considerably endangered by drift effects if random mating prevails. The recessive gene may reach the same effectivity in establishment as a dominant gene, only if the recessive homozygote mates exclusively with its own type during the period of establishment.