蛋白质组学在绵羊育种中的应用

蛋白质组学作为后基因组时代的主要研究方向和重要研究手段之一,近年来被广泛地应用于生物科学及交叉学科的各个领域.蛋白质组学还衍生出一套在整体水平上或生物动态水平上研究蛋白质及其之间相互作用的方法.日渐成熟的蛋白质组学技术极大地推动了蛋白质组学的研究进程.这些技术不仅包括蛋白质提取和分离,还包括对目的蛋白质进行鉴定和分析,这为蛋白质组学研究提供了技术支持.文章简述了蛋白质组学的基本方法、技术路线,并重点阐述了蛋白质组学在绵羊育种研究中的应用情况.     

蛋白质组学研究与应用

蛋白质组学是继基因组学之后20世纪末兴起的前沿学科热点．阐述了蛋白质组的概念、蛋白质组学研究的内容与特点和研究意义；综述了蛋白质纽学研究中蛋白质组样品的制备方法、所运用的蛋白质组分离技术、鉴定技术等研究技术手段及其特点，以及蛋白质组学在基础生物学、基础医学、疾病诊断与治疗、新药研究开发中的应用现状与前景；介绍了国内、外蛋白质组学研究领域的最新进展．     

质膜及其微区的蛋白质组学研究进展

质膜蛋白质由于低丰度和强疏水性等特点,给其提取、溶解、分离以及鉴定带来了很大的困难,因此使质膜蛋白质组学研究成为蛋白质组学研究中的难点和热点.以哺乳动物细胞质膜为例,主要从质膜及其微区的富集、溶解、分离、鉴定方法等方面介绍了细胞质膜及微区蛋白质组以及复合物的研究进展.     

蛋白质组学的研究进展

蛋白质组学是近几年出现的生物学者研究的热点和新领域.人类基因组"工作框架图"的完成,标志着后基 因组时代的来临,在后基因组时代,研究的中心将从揭示生命的所有遗传信息转移到在整体水平上对功能的研究,主 要以蛋白质组为中心阐述了一些与其相关的概念、研究内容、技术及其应用方面的研究进展;在这其中,蛋白质组学 的相关概念包括结构基因组学和功能蛋白质组学;而研究技术主要有2-DE技术、生物质谱技术、蛋白质芯片技术以及 生物信息学技术等.     

差异蛋白质组学及其应用

蛋白质组学是后基因组研究中最主要的部分,其研究策略有2种:一种是"完全"蛋白质组学,目的是检测一种细胞类型或一种组织内基因组表达的所有蛋白质; 另一种是"差异"蛋白质组学,主要是筛选和鉴定不同种类或状态下各样本之间蛋白质组的区别与变化.着重介绍了差异蛋白质组学的特点、研究内容和应用前景,简要介绍了适用于差异蛋白质组学研究的相关技术和近期发展概况.     

蛋白质与蛋白质组学的研究进展

介绍了蛋白质、蛋白质组和蛋白质组学的基本概念，蛋白质组学的研究内容、方法技术、形成与发展。     

蛋白质组学及其在神经科学研究中的应用

蛋白质组学是后基因组时代研究中兴起还不到十年的新型学科,但已应用到生命科学与基础医学研究中的各个领域.对蛋白质组进行大规模的分析可使我们更加深入了解生物体的结构与功能.在神经科学研究领域,对于神经突触、受体复合物以及其他的神经元和神经胶质特性的研究因为该学科的渗透,近年来取得了辉煌成就.然而,蛋白质组学相关技术用于神经系统的研究还面临着很多挑战,如脑蛋白样品的准备、脑的复杂性、有限的数据库资源和生物信息学工具等.蛋白质组学技术与其他功能基因组学研究方法,如基因芯片、网络生物学分析技术等相结合,将在基因及其功能、神经生理学与病理学之间架起一道重要的桥梁.     

蛋白质组学及其在肿瘤研究中的应用

蛋白质组学是在细胞的整体蛋白质水平上进行研究、从蛋白质整体活动的角度来认识生命活动规律的一门新学科.由于生命的表现形式最终是由蛋白质决定的,因此,从蛋白质组的角度研究肿瘤的诊断与治疗具有积极的现实意义.     

基因组学与蛋白质组学

介绍了基因组与基因组学、蛋白质与蛋白质组学的概念及功能基因组学和蛋白质组学的研究方法。     

蛋白质组学用于疾病特殊蛋白质的鉴定

蛋白质组学是一门新的启动技术.它将便利越过一切生物学系统或疾病的蛋白质系统分析.在疾病特殊蛋白质的鉴别过程蛋白质组学高度补充基因组方法,且第一次提供科学家使蛋白质组信息,表达mRNA,它们各自的蛋白质和亚细胞定位一体化的能力.期望会导致对疾病机制新的重大认识.     

溶藻弧菌对水产动物致病性及其防治的研究进展

溶藻弧菌是一种常见的海洋伺机性病原体,是引起人类、水生动物细菌性疾病的重要病原之一。与其它病原微生物一样,弧菌的致病性与其产生的多种毒力因子息息相关,毒力基因是产生各种毒力因子的物质基础。本文对溶藻弧菌的病原学、致病机理及其防治等方面的最新研究成果进行了综述。     

Complex lipid determines tissue-specific replication of Mycobacterium tuberculosis in mice

Tuberculosis is the leading cause of death in the world resulting from a single bacterial infection. Despite its enormous burden on world health, little is known about the molecular mechanisms of pathogenesis of Mycobacterium tuberculosis. Bacterial multiplication and concomitant tissue damage within an infected host, including experimentally infected mice, occurs primarily in the lungs-the favoured niche of M. tuberculosis. Although it has been proposed that the distinctive cell wall of M. tuberculosis is important for virulence, rigorous genetic proof has been lacking. Here, using signature-tagged mutagenesis, we isolated three attenuated M. tuberculosis mutants that cannot synthesize or transport a complex, cell wall-associated lipid called phthiocerol dimycocerosate (PDIM) which is found only in pathogenic mycobacteria. Two mutants have transposon insertions affecting genes implicated in PDIM synthesis; the third has a disruption in a gene encoding a large transmembrane protein required for proper subcellular localization of PDIM. Synthesis and transport of this complex lipid is only required for growth in the lung; all three mutants are unaffected for growth in the liver and spleen. This clearly shows that a lipid is required for M. tuberculosis virulence.     

羊布鲁菌外膜蛋白Omp22的原核表达及鉴定

布鲁菌外膜蛋白Omp22与布鲁菌致病性相关,又可诱导机体免疫反应。为进一步研究Omp22的特性,对其进行原核表达,并初步鉴定。根据Genbank序列,设计并合成羊布鲁菌外膜蛋白Omp22引物。以羊布鲁菌M5株基因组DAN为模板,PCR扩增并克隆入原核表达载体pGEX-4T-1中。酶切鉴定正确后,测定序列;将重组质粒pGEX-4T-1-Omp22电转化大肠杆菌DH5α中,诱导表达,并采用羊布鲁菌兔免疫血清,Western-blot初步检测融合蛋白GST-Omp22表达情况和免疫学特性。结果PCR扩增出约660bp目的条带,与预期相符;酶切鉴定和测序结果表明,成功构建了pGEX-4T-1-Omp22原核表达载体;优化诱导温度,结果表明,在25℃诱导表达时,该融合蛋白大部分出现在上清中;Western-blot结果证实,GST-Omp22可与羊布鲁菌兔免疫血清特异性结合。说明成功构建了Omp22蛋白原核表达载体并进行了可溶性表达;该融合蛋白可与羊布鲁菌兔免疫血清特异性结合。Omp22蛋白的表达,为进一步研究布鲁菌的致病机制以及疫苗研制奠定基础。     

H5N1的结构与致病机制

以禽流感病毒H5N1的两种基质蛋白HA、NA的结构为基础,分析了H5N1的基因结构,探索其毒力基础和致病的分子机制.     

Maintenance of an unfolded polypeptide by a cognate chaperone in bacterial type III secretion.

Many bacterial pathogens use a type III protein secretion system to deliver virulence effector proteins directly into the host cell cytosol, where they modulate cellular processes. A requirement for the effective translocation of several such effector proteins is the binding of specific cytosolic chaperones, which typically interact with discrete domains in the virulence factors. We report here the crystal structure at 1.9 A resolution of the chaperone-binding domain of the Salmonella effector protein SptP with its cognate chaperone SicP. The structure reveals that this domain is maintained in an extended, unfolded conformation that is wound around three successive chaperone molecules. Short segments from two different SptP molecules are juxtaposed by the chaperones, where they dimerize across a hydrophobic interface. These results imply that the chaperones associated with the type III secretion system maintain their substrates in a secretion-competent state that is capable of engaging the secretion machinery to travel through the type III apparatus in an unfolded or partially folded manner.     

Exploitation of syndecan-1 shedding by Pseudomonas aeruginosa enhances virulence

Cell-surface heparan sulphate proteoglycans (HSPGs) are ubiquitous and abundant receptors/co-receptors of extracellular ligands, including many microbes. Their role in microbial infections is poorly defined, however, because no cell-surface HSPG has been clearly connected to the pathogenesis of a particular microbe. We have previously shown that Pseudomonas aeruginosa, through its virulence factor LasA, enhances the in vitro shedding of syndecan-1-the predominant cell-surface HSPG of epithelia. Here we show that shedding of syndecan-1 is also activated by P. aeruginosa in vivo, and that the resulting syndecan-1 ectodomains enhance bacterial virulence in newborn mice. Newborn mice deficient in syndecan-1 resist P. aeruginosa lung infection but become susceptible when given purified syndecan-1 ectodomains or heparin, but not when given ectodomain core protein, indicating that the ectodomain's heparan sulphate chains are the effectors. In wild-type newborn mice, inhibition of syndecan-1 shedding or inactivation of the shed ectodomain's heparan sulphate chains prevents lung infection. Our findings uncover a pathogenetic mechanism in which a host response to tissue injury-syndecan-1 shedding-is exploited to enhance microbial virulence apparently by modulating host defences.     

疏肝解毒法治疗病毒性肝炎

本文运用中医辨证论治的方法,从分析病毒性肝炎的病因病机入手,提出了用疏肝解毒法治疗病毒性肝炎,并以实例证明此法的可行性和可靠性,及其在治疗病毒性肝炎中有着广泛应用价值。     

Chaperone release and unfolding of substrates in type III secretion

Type III protein secretion systems are essential virulence factors of many bacteria pathogenic to humans, animals and plants. These systems mediate the transfer of bacterial virulence proteins directly into the host cell cytoplasm. Proteins are thought to travel this pathway in a largely unfolded manner, and a family of customized cytoplasmic chaperones, which specifically bind cognate secreted proteins, are essential for secretion. Here we show that InvC, an ATPase associated with a Salmonella enterica type III secretion system, has a critical function in substrate recognition. Furthermore, InvC induces chaperone release from and unfolding of the cognate secreted protein in an ATP-dependent manner. Our results show a similarity between the mechanisms of substrate recognition by type III protein secretion systems and AAA + ATPase disassembly machines.     

Thymidine kinase gene mutation leads to reduced virulence of pseudorabies virus

To explore correlation between the tk gene structure of pseudorabies virus (PRV) and its virulence, to study the effect of the gene mutation on PRV biological properties, and to investigate mechinism of reduced virulence, thymidine kinase (TK)-deficient mutant of pseudorabies virus strain Hubei (PRV HB) was isolated by selection for resistance to 5-bromodeoxyuridine. The tk genes of PRV HB and its TK^― mutant were cloned and sequenced. 1587 base pairs of the tk gene and flanking regions of wild-type (wt) virus were sequenced, which included an open reading frame (ORF) of 1098 bp encoding a protein of 366 amino acids. The ORF contained two 137-bp repeated sequences, which were connected by an adenosine. 1458 bp of the tk and flanking regions of TK^― mutant were sequenced. Analysis of the tk gene sequence of TK^― mutant indicated that one of 137 bp repeated sequence and the connecting adenosine in the tk gene of the wt virus was deleted and a repeated sequence of 8 nucleotides (GCGCGCC) was inserted. All other nucleotides of TK^―mutant were identical to that of wt virus. Deletion and insertion of the nucleotide sequence resulted in a frameshift and a premature chain termination, and the resultant TK protein was not active. Analysis of the amino acid sequence revealed that TK protein of PRV HB contained the conserved consensus sequence of herpesviral TKs and an additional conserved-DHR-motif. The results of this work also indicated that TK^― mutant was genetically stable. Compared to PRV HB, virulence of TK^― mutant was greatly decreased. Mice vaccinated with TK^― mutant were completely protected against a lethal challenge with virulent PRV (HB).     

Increased virulence of Yersinia pseudotuberculosis by two independent mutations

A chromosomally encoded protein, which mediates invasion into HeLa cells was recently identified in Yersinia pseudotuberculosis. The role of this protein (invasin) in the virulence process was not, however, investigated. We show that mutation of the invasin gene in Y. pseudotuberculosis abolishes the ability of the bacteria to invade HeLa cells. When mice were challenged by intraperitoneal injection both the mutant and the wild-type strain produced infections of similar virulence but mutant showed a slower rate of infection after oral challenge. A double mutant, carrying an additional mutation in the gene coding for the Yop1 protein, was also constructed. The double mutant was significantly more virulent than either the wild-type or the corresponding single mutants. Y. pestis, in contrast to Y. pseudotuberculosis lacks the ability to express either invasin or Yop1, sequence analysis of the yopA gene from both Y. pestis and Y. pseudotuberculosis shows that the yopA gene of Y. pestis contains a point-mutation leading to a reading-frame shift. When the yopA+ gene was introduced into Y. pestis the virulence of this strain was reduced. These results may provide insight into the rise and fall of plague epidemics caused by Y. pestis.