病毒装配的分子生物学

病毒的装配是病毒复制和增殖过程中的一个重要步骤,它不是一个简单的静态的结构生物学问题,而是一个综合的动态的过程。它至少包括了下列几个问题：１．病毒的衣壳蛋白亚基是如何互相识别和装配成病毒衣壳的,是什么机制来控制这一过程的？２．病毒的基因组（ＤＮＡ或Ｒ...     

免出血症病毒的形态与结构的研究

免出血症病毒无锡株A_2R—3的完整病毒颗粒呈球形,廿面体等轴对称、无囊膜、其直径约为33—37nm。空心病毒粒子亦可看到,其核心直径为21—25nm。病毒衣壳包裹在颗粒的最外层,由紧密连结的子粒所组成,子粒排列规则,呈管状结构。其长度约为5—6nm,中心孔径为2—3nm。廿面体对称的等边三角形的面由6个子粒所构成,即每条边上排列3个子粒。据此推算出病毒衣壳的子粒总数为42,分负数为4,三角形面数为80,结构单位数为240。     

一种含有单链RNA的香菇球状病毒

从生长不正常的香菇(Lentinus edodes(Berk.)Sing)菌株中分离到一种等轴对称含单链RNA的病毒颗粒。病毒颗粒在电镜下直径为33～34nm,在SDS-聚丙烯酰胺凝胶电泳中病毒外壳蛋白分子量为22000道尔顿。病毒核酸径DNase1和SI酶解试验及热变性紫外吸收曲线试验证明为单链RNA,在1.5％的琼脂糖凝胶电泳中,病毒核酸呈现一条带,分子量为2.38×10~6道尔顿。     

真核藻类的病毒和病毒类粒子（VLPs）

真核藻类的病毒和病毒类粒子（ＶＬＰｓ）赵以军石正丽（中国科学院水生生物研究所,武汉４３００７２）（中国科学院武汉病毒研究所,武汉４３００７１）ＶｉｒｕｓｅｓａｎｄＶｉｒｕｓ－ｌｉｋｅＰｅａｒｔｉｃｌｅｓｏｆＥｕｋａｒｙｏｔｉｃＡｌｇａｅＺｈａｏＹｉｊ...     

提高滴定病毒准确度的方法：胡宁病毒蚀斑检测的标准化

胡宁病毒是沙状病毒族中的一员,它可引起阿根廷出血热。阿根廷出血热是以血液、心血管、神经和肾脏改变为特征的。阿根廷出血热的诊断通常是通过全血接种乳鼠,该方法可获得经血清学证实病例中的50～70％查到胡宁病毒。Amtrosia等企图用单层细胞分离病毒,阳性标本的检出率却很低。胡宁病毒在单层细胞中明显缺乏敏感性的原因还不清楚。     

TT病毒的分类及致病性研究

TT病毒是近年来新发现的一种肝炎相关病毒,隶属圆环病毒科,迄今已确认了至少28种基因型。目前,人们对TT病毒的致病性存在很大的争议,它对肝脏确切的致病作用尚待进一步研究证实。最近的研究提示,TT病毒与呼吸系统疾病有一定相关性,其开放阅读框架产物可损害肾上皮细胞。     

Immunochemical analysis of the products of translation of mRNA hybridized with the left HindIII-A-Sa1-fragment of vaccinia virus DNA

The left HindIII-A-Sal fragment of the vaccinia virus DNA has been analyzed by the technique of mRNA hybridizational selection with the subsequent translation in cell-free protein-synthesizing system from the rabbit reticulocytes. The viral mRNA hybridizable with the fragment was shown to direct the synthesis of 12, 17, 27, 42, 70 kD polypeptides in the cell-free protein-synthesizing system. Each of 12 and 42 kD polypeptides was demonstrated to react specifically with antisera to structural p12 and p42 coat proteins. The structural coat proteins p12, p20, p42 of the vaccinia virus are concluded to be the products of the same viral gene.     

Subcellular distribution of viral structural proteins during simian virus 40 infection.

The amounts of simian virus 40 structural polypeptides Vp1, Vp2, and Vp3 in different subcellular fractions at various times after lytic infection were determined by a quantitative immunoblotting procedure. Simian virus 40-infected cells were lysed with a buffer containing Nonidet P-40 to yield a soluble fraction. The Nonidet P-40-insoluble fraction was further fractionated in the presence of deoxycholate and Tween 40 to yield a soluble fraction (cytoskeletal) and an insoluble fraction (Nuc), which is primarily cell nuclei. At 33 h postinfection, the majority of viral structural proteins was found in the cell nucleus, whereas, at 48 to 65 h postinfection, Vp1 was distributed evenly among all cell fractions and Vp2 and Vp3 were found predominantly in the cytoskeletal and Nuc fractions. Thus, not all of the viral polypeptides synthesized in the cytoplasm migrated into the cell nucleus. Throughout infection, the molar ratio (Vp3/Vp2) was rather constant in all subcellular fractions, indicating that the synthesis or processing or both of Vp2 and Vp3 are coordinately regulated. The molar ratio of Vp1/(Vp2 + Vp3) varied among the fractions. The Vp1/(Vp2 + Vp3) molar ratio in the soluble fraction varied during the course of infection; however, constant ratios were maintained in the cytoskeletal and Nuc fractions. Thus, the mechanism which controls the movement of Vp1 to different compartments of the cell appears to be different from that of Vp2 and Vp3. The Vp1/(Vp2 + Vp3) value in the Nuc fraction was similar to the ratio found in virus particles. The constant molar distribution of Vp1, Vp2, and Vp3 in the Nuc fraction throughout infection suggests that there is a specific mechanism which regulates the transport of viral structural proteins. These results support the hypothesis that the structural proteins of simian virus 40 are transported into the cell nucleus in precise proportions.     

小鹅瘟病毒纯化及其理化特性的研究

详细描述了小鹅瘟病毒(Goslin—plague virus,GPV)扬州株的浓缩和纯化过程,并论述了其理化特性。试验证明,GPV在氯化铯中主要病毒区带的浮密度为11.31～1.35g/ml,电镜下可见空壳和实心两种病毒粒子,大小20～22nm,沉降系数90.5S。用Sepharose 4B柱层析纯化的病毒,等电点为4.3。GPV有3种结构多肽,即Vp1、Vp2和Vp3,分子量分别为85 000,61 000,57 500道尔顿,其中Vp3为主要结构多肽,纯化的病毒粒子能使鹅胚致死。     

草鱼出血病病毒多肽的免疫原性

采用中和试验比较了草鱼出血病病毒ＧＣＨＶ８７３的１１个多肽的免疫原性,确定了主要中和抗原。纯化的单个病毒多肽免疫家兔获得抗血清,多太ＶＰ１、ＶＰ５、ＶＰ６和ＶＰ９的抗血清具有中和效价,而ＶＰ２、ＶＰ３、ＶＰ４、ＶＰ８、ＶＰ１０和ＶＰ１１则不能诱生中和抗体。其中ＶＰ５的抗血清中和效价最高,因此,ＶＰ６极可能是病毒多肽中的主要中和抗原。     

In vitro morphogenesis of foot-and-mouth disease virus.

Foot-and-mouth disease virion RNA is translated efficiently and completely in a rabbit reticulocyte lysate cell-free system. Treatment of cell-free lysates with monospecific serum prepared against the individual viral structural proteins or with monoclonal antibodies prepared against the inactivated virus or against a viral structural protein precipitated all of the structural proteins, suggesting that structural protein complexes were formed in vitro. Sucrose gradient analysis of the cell-free lysate indicated that complexes sedimenting at 5, 14, 60 to 70, and ca. 110S were assembled in vitro. Structural proteins VP0, VP1, and VP3 were the major polypeptides found in these complexes. The material sedimenting at 110S, i.e., containing VP0, VP1, and VP3, was precipitated by a 140S-specific monoclonal antibody but not by a 12S subunit-specific monoclonal antibody, suggesting that this capsid structure contained at least one epitope present on the intact virus.     

The errors in assembly of MuLV in interferon treated cells

Interferon treatment of JLSV-6 cells chronically infected with Rauscher MuLV leads to the formation of noninfectious particles (interferon virions) containing the structural proteins of env and gag genes as well as additional viral polypeptides. In the control virions the major glycoprotein detected is gp71, interferon virions contain in addition to gp71 and 85k dalton (gp85) glucosamine-containing, fucose-deficient glycoprotein which is recognized by antiserum to MuLV but not by the gp71 antiserum. The surface iodination of the intact virions indicates that both gp71 and gp85 are the major components of the external virions envelope. However, unlike the control virions in which gp71 associates with p15E (gp90), the gp71-p15E complex was not detected in interferon virions. The analysis of the iodinated proteins of the disrupted interferon virions revealed the presence of 85k and 65k dalton polypeptides preciptable with antiserum against MuLV, which are not present in the control virions. The difference in the polypeptide pattern of virions produced in the presence of interferon does not seem to be a consequence of the slowdown in the synthesis of viral proteins or their processing in the interferon-treated cells. Both the structural proteins of env and gag genes seem to be synthesized and processed at a comparable rate in the interferon-treated and -untreated cells. These results indicate an alteration of virus assembly in the presence of interferon.     

Translation of three mouse hepatitis virus strain A59 subgenomic RNAs in Xenopus laevis oocytes

We have purified the seven virus-specific RNAs which were previously shown to be induced in Sac(-) cells upon infection with mouse hepatitis virus strain A59 (W. J. M. Spaan, P. J. M. Rottier, M. C. Horzinek, and B. A. M. van der Zeijst, Virology 108:424-434, 1981). The individual RNAs, prepared by agarose gel electrophoresis of the polyadenylated RNA fraction from infected cells, were obtained pure, except for the preparations of RNAs 4, 5, and 6, which contained some contamination of RNA 7. The RNAs were microinjected into Xenopus laevis oocytes, and after incubation of these cells in the presence of [35S]methionine, the proteins synthesized were analyzed by polyacrylamide gel electrophoresis. Whereas no translation products of RNAs 1, 2, 4, and 5 were detected, the synthesis of virus-specific polypeptides coded by RNAs 3, 6, and 7 was observed. RNA 7 (0.6 X 10(6) daltons) directed the synthesis of a 54,000-molecular-weight polypeptide which comigrated with viral nucleocapsid protein and which was immunoprecipitated by antiserum from mice that had been infected with the virus. RNA 6 (0.9 X 10(6) daltons) directed the synthesis of three polypeptides with molecular weights of 24,000, 25,500, and 26,500, which migrated with the same electrophoretic mobilities as three low-molecular-weight virion polypeptides. After injection of RNA 3 (3.0 X 10(6) daltons), a polypeptide with a molecular weight of about 150,000 was immunoprecipitated. This polypeptide had no counterpart in the virion, but comigrated with a virus-specific glycoprotein present in infected cells which is immunoprecipitated by a rabbit antiserum against the mouse hepatitis virus strain A59 structural proteins. This antiserum could also immunoprecipitate the translation products of RNAs 3, 6, and 7. These results indicate that RNAs 3, 6, and 7 encode viral structural proteins. The significance of the data with respect to the strategy of coronavirus replication is discussed.     

Serological Analysis of the Deoxyribonucleic Acid Polymerase of Avian Oncornaviruses II. Comparison of Avian Deoxyribonucleic Acid Polymerases

Monospecific antiserum prepared against the isolated deoxyribonucleic acid (DNA) polymerase of avian myeloblastosis virus (AMV) neutralized the endogenous ribonucleic acid-instructed DNA polymerase activity of detergent-disrupted virus. The viral polymerase was serologically unrelated to the seven major structural polypeptides of AMV. Furthermore, the viral enzyme was distinguished from normal cellular DNA polymerases by serological criteria; thus, antiserum against the viral enzyme neutralized its homologous antigen but not normal cellular DNA polymerases. Neutralization by antibody of viral DNA polymerase activity was observed with all avian leukemia-sarcoma viruses tested, irrespective of viral antigenic subtype. The DNA polymerase activity of avian reticuloendotheliosis virus, and of a variety of mammalian oncornaviruses, was not neutralized by antisera against the AMV polymerase. Immunological analysis of the RSValpha(O) mutant, which is deficient in DNA polymerase activity, shows this mutant to lack demonstrable polymerase antigen. Viral polymerase was identified by immunofluorescence as a cytoplasmic constituent in virus-producing chicken cells; polymerase antigen was not detected in uninfected (gs(-)) chicken cells.     

Immunological characterization of rapeseed myrosinase

A purified 75-kDa myrosinase and a crude rapeseed myrosinase fraction were used as antigens to produce mouse anti-myrosinase monoclonal antibodies. The 75-kDa myrosinase was also used to produce a polyclonal rabbit antiserum. The antiserum and one monoclonal antibody reacted with three distinct rapeseed polypeptides of 75, 70 and 65 kDa (M75, M70 and M65, respectively). A second set of monoclonal antibodies reacted exclusively with the 75-kDa form of myrosinase, and a third set showed specificity towards two components of 52 and 50 kDa (myrosinase-binding proteins, MBP52 and MBP50, respectively). MBP52 and MBP50 lack inherent myrosinase activity, but are nevertheless capable of mediating immunoprecipitation of myrosinase due to their interaction with myrosinase. Gel chromatography and glycerol gradient centrifugation experiments resolved two myrosinase-containing fractions. One of these had an approximate molecular mass of 140 kDa and consisted of disulfide-linked dimers of the 75-kDa myrosinase. The other fraction was heterogeneous in size with molecular masses ranging from 250 kDa to approximately 1 MDa. The high-molecular-mass fractions contained complexes consisting of disulfide-linked 70-kDa and 65-kDa myrosinases and non-covalently bound 52-kDa and 50-kDa myrosinase-binding proteins.