炎症及炎症耐受模型筛选广谱趋化因子抑制肽

通过减少炎性组织或细胞趋化因子及炎性因子的表达量能将炎症性病理过程抑制在起始阶段．我们通过体外构建人外周血单个核细胞LPS激活的急性炎症模型及内毒素耐受模型,进行噬菌体肽库亲和筛选,ELISA检测与炎性PBMC的结合能力,分泌抑制实验筛选抑制性噬菌体克隆,经趋化抑制、竞争结合及生成抑制实验检测体外活性,大鼠足肿胀及关节炎模型检验多肽体内作用,SqRT-PCR检测趋化因子及TTP的mRNA水平,探讨其作用机制．筛选到的目标多肽CI-S5趋化抑制率达到48.72%,明显抑制噬菌体阳性克隆P15与LPS激活PBMC的结合,动物试验能明显降低大鼠足肿胀及关节炎症．机制研究显示,CI-S5多肽能降低3种趋化因子的表达量,并调节TTP使其表达增加,提示CI-S5能够靶向炎症前期PBMC,为炎症治疗提供了针对早期急性炎症反应的广谱小分子抑制肽．     

趋化因子受体 CCR5 亲合短肽的筛选

趋化因子受体 5 (CCR5) 是 HIV-1 与宿主细胞结合的辅助因子之一,其功能缺失或被 CCR5 拮抗剂封闭则会阻止 HIV-1 感染细胞 . 为得到与 CCR5 特异结合的肽类拮抗剂,采用噬菌体展示技术,以稳定表达 CCR5 的 CHO 细胞 (CHO/CCR5) 作为靶标,通过噬菌体随机 12 肽库筛选与 CCR5 特异结合的多肽;经过四轮筛选后,挑选 20 个阳性噬菌体克隆进行测序,从中得到 11 个含有 AFDWTFVPSLIL 序列的小分子肽 . 含该序列的噬菌体能与抗人 CCR5 单抗 (2D7) 竞争性结合 CCR5 ,且合成肽 AFDWTFVPSLIL 对趋化因子 RANTES 与 CHO/CCR5 的结合具有明显的抑制作用,初步证明该小肽与 CCR5 具有特异性结合作用 .     

以表位模拟肽为亲和靶标的肉毒毒素受体结合抑制剂的筛选及鉴定

以设计合成的A型肉毒毒素表位模拟肽为亲和靶标,对噬菌体随机肽库进行筛选,寻找能与A型肉毒毒素表位模拟肽特异结合并能拮抗毒素毒性效应的分子,通过ELISA鉴定阳性克隆,并对鉴定的阳性克隆进行特异性分析及DNA测序。氨基酸序列同源性分析发现,针对P4、P5表位模拟肽获得了两条特异的结合序列,并通过动物保护实验在噬菌体展示肽水平对特异结合分子的毒素毒性拮抗效应进行了初步研究,初步证明结合肽对A型肉毒毒素有一定的保护作用。     

利用噬菌体展示肽库筛选鸡传染性支气管炎病毒模拟抗原表位

目的：利用噬菌体展示肽库技术筛选鸡传染性支气管炎病毒（IBV）的模拟抗原表位。方法：用IBV阳性血清纯化IgG作为靶标。对噬菌体展示随机12肽库进行筛选，通过ELISA和竞争抑制ELISA鉴定筛选克隆的结合特性，并对阳性克隆提取ssDNA进行测序分析。结果：3轮生物淘洗后，目标噬菌体得到125倍富集。随机挑选50个克隆进行ELISA和竞争抑制ELISA。其中有12个噬菌体克隆可以与IBV阳性血清高特异性结合。测序分析发现，这12个克隆带有2种氨基酸序列。即KSPKHSSSALHF和SFFQLNLHRPTS。且未发现这2种序列与GenBank中已发表的IBV氨基酸序列有同源性。结论：结果提示。这2个肽可能是IBV抗原的模拟表位。     

噬菌体展示随机十肽库的构建及抗WSSV单链抗体A1的模拟抗原表位的淘选和鉴定

本实验以pCANTAB5E噬菌粒为载体,成功构建了较高容量的噬菌体展示随机十肽库,并将其应用于抗原模拟表位的淘选和鉴定。将一种特异识别对虾白斑综合症病毒(Whitespotsyndromevirus,WSSV)的单链抗体A1对十肽库和十五肽库分别进行淘选,结果得到一系列能与单链抗体A1特异性结合的阳性克隆。将这些阳性克隆所编码的多肽氨基酸序列与已知的单链抗体A1的抗原WSSV388片段氨基酸序列做比对,发现多数阳性多肽序列都与WSSV388片段序列的C端一处K????R??R?QS的氨基酸片段相似,由此推论单链抗体A1的模拟抗原表位可能是由该不连续氨基酸片段所构成的构象表位,而非线性表位。研究结果表明,噬菌体展示随机肽库技术是一种用于研究抗原表位结构的有效方法,有助于进一步探讨WSSV的结构蛋白的构象及功能,以及相应单链抗体与细胞受体相互作用的机理。     

从噬菌体展示肽库中筛选脂质A的结合肽

以脂质A为靶标,筛选噬菌体展示十二肽库,三轮后随机挑取14个噬菌体克隆进行结合活性鉴定,并对5个克隆进行序列分析。结果表明,14个克隆全部是阳性克隆,测序结果显示4个阳性克隆的序列完全一样。说明筛选到了一个脂质A的结合多肽。     

从噬菌体随机肽库中筛选流感病毒神经氨酸酶的抑制多肽

目的:从噬菌体呈现12肽库中筛选与流感病毒神经氨酸酶特异性结合的肽。方法:以甲三型流感病毒裂解疫苗原液为靶分子,经过3轮生物淘选,从噬菌体随机肽库中筛选与之结合的噬菌体。用ELISA方法鉴定噬菌体克隆与靶分子的结合力,用荧光方法测定噬菌体克隆对流感病毒A/Sydney/5/97(H3N2)神经氨酸酶的抑制活性。对筛选到的阳性克隆进行DNA序列测定并推导出相应的氨基酸序列。结果:经过3轮筛选后,42个噬菌体克隆与靶分子有高度亲和力,23个噬菌体克隆对流感病毒A/Sydney/5/97(H3N2)神经氨酸酶有抑制活性。对27个噬菌体克隆的测序结果表明,分别有10个和2个克隆的序列是一致的,其氨基酸序列分别为KSLSRHDHIHHH和WPRHHHSASVQT。结论:通过噬菌体肽库筛选到抑制流感病毒神经氨酸酶的12肽,为进一步研究对流感病毒神经氨酸酶有抑制活性的分子药物奠定了基础。     

应用噬菌体随机肽库技术筛选丙肝病毒NS3抗原模拟表位

以抗 HCVNS3的单克隆抗体作为固相筛选分子 ,对人工合成的噬菌体随机 12肽库进行 5轮“吸附 洗脱 扩增”的筛选过程 ,随机挑取 4 2个克隆 ,经噬菌体酶联免疫吸附法 (ELISA)鉴定并进行交叉反应实验以及竞争抑制性结合实验 ,最后对所选克隆进行DNA序列分析 ,以确定HCVNS3抗原的模拟表位。经噬菌体富集后 ,从随机筛选的 4 2个克隆中得到 11个阳性克隆 ,确定氨基酸序列XXIXXXXMSNXX为HCVNS3的模拟表位。我们用噬菌体12肽库成功筛选得到HCVNS3的模拟表位 ,为开展用HCV模拟表位探索HCV的防治研究创造了条件     

表位九肽库的构建及人Ⅳ型胶原酶特异结合肽的筛选

将人工合成的编码九肽的随机序列DNA片段克隆进丝状噬菌体表达载体FUSE5,经多次电击转化和表达,获得肽段与噬菌体pⅢ蛋白融合并展示在噬菌体表面的随机序列九肽表位肽库。库容量达10 10个克隆。以Ⅳ型胶原酶为靶蛋白,采用亲和纯化筛选模式,从中筛选出Ⅳ型胶原酶结合肽。进一步ELISA检测筛选出与Ⅳ型胶原酶特异结合的20个阳性克隆。序列分析发现一组肽含有WDXXD的共同序列,一组含有WVGXXR的共同序列。其中WDXXD的序列与Ⅳ型胶原酶单链抗体可变区序列同源。结果表明,多肽库是筛选蛋白特异结合肽的有力工具,表位九肽库的构建和筛选方法的建立为进一步应用筛选具有高亲和力的特异结合肽奠定了基础。     

噬菌体展示随机十肽库的构建及抗WSSV单链抗体A1的模拟抗原表位的淘选和鉴定

本实验以pCANTAB 5 E噬菌粒为载体,成功构建了较高容量的噬菌体展示随机十肽库,并将其应用于抗原模拟表位的淘选和鉴定.将一种特异识别对虾白斑综合症病毒(White spot syndrome virus, WSSV)的单链抗体A1对十肽库和十五肽库分别进行淘选,结果得到一系列能与单链抗体A1特异性结合的阳性克隆.将这些阳性克隆所编码的多肽氨基酸序列与已知的单链抗体A1的抗原WSSV388片段氨基酸序列做比对,发现多数阳性多肽序列都与WSSV388片段序列的C端一处K····R··R·QS的氨基酸片段相似,由此推论单链抗体A1的模拟抗原表位可能是由该不连续氨基酸片段所构成的构象表位,而非线性表位.研究结果表明,噬菌体展示随机肽库技术是一种用于研究抗原表位结构的有效方法,有助于进一步探讨WSSV的结构蛋白的构象及功能,以及相应单链抗体与细胞受体相互作用的机理.     

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.     

A Unique Protease Cleavage Site Predicted in the Spike Protein of the Novel Pneumonia Coronavirus (2019-nCoV) Potentially Related to Viral Transmissibility

正Dear Editor,In December 2019, a novel human coronavirus caused an epidemic of severe pneumonia(Coronavirus Disease 2019,COVID-19) in Wuhan, Hubei, China(Wu et al. 2020; Zhu et al. 2020). So far, this virus has spread to all areas of China and even to other countries. The epidemic has caused 67,102 confirmed infections with 1526 fatal cases     

Curcuminoids as inhibitors of thioredoxin reductase: A receptor based pharmacophore study with distance mapping of the active site

Curcumin is the yellow pigment of turmeric that interacts irreversibly forming an adduct with thioredoxin reductase (TrxR), an enzyme responsible for redox control of cell and defence against oxidative stress. Docking at both the active sites of TrxR was performed to compare the potency of three naturally occurring curcuminoids, namely curcumin, demethoxy curcumin and bis-demethoxy curcumin. Results show that active sites of TrxR occur at the junction of E and F chains. Volume and area of both cavities is predicted. It has been concluded by distance mapping of the most active conformations that Se atom of catalytic residue SeCYS498, is at a distance of 3.56 from C13 of demethoxy curcumin at the E chain active site, whereas C13 carbon atom forms adduct with Se atom of SeCys 498. We report that at least one methoxy group in curcuminoids is necessary for interation with catalytic residues of thioredoxin. Pharmacophore of both active sites of the TrxR receptor for curcumin and demethoxy curcumin molecules has been drawn and proposed for design and synthesis of most probable potent antiproliferative synthetic drugs.     

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.