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1.
Anatomy of the herpes simplex virus 1 strain F glycoprotein B gene: primary sequence and predicted protein structure of the wild type and of monoclonal antibody-resistant mutants. 总被引:58,自引:49,他引:9
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In this paper we report the nucleotide sequence and predicted amino acid sequence of glycoprotein B of herpes simplex virus 1 strain F and the amino acid substitutions in the domains of the glycoprotein B gene of three mutants selected for resistance to monoclonal antibody H126-5 or H233 but not to both. Analyses of the amino acid sequence with respect to hydropathicity and secondary structure yielded a two-dimensional model of the protein. The model predicts an N-terminal, 29-amino-acid cleavable signal sequence, a 696-amino-acid hydrophilic surface domain containing six potential sites for N-linked glycosylation, a 69-amino-acid hydrophobic domain containing three segments traversing the membrane, and a charged 109-amino-acid domain projecting into the cytoplasm and previously shown to marker rescue glycoprotein B syn mutations. The nucleotide sequence of the mutant glycoprotein B DNA fragments previously shown to marker transfer or rescue the mutations revealed that the amino acid substitutions cluster in the hydrophilic surface domain between amino acids 273 and 305. Analyses of the secondary structure of these regions, coupled with the experimentally derived observation that the H126-5- and H233-antibody cognitive sites do not overlap, indicate the approximate locations of the epitopes of these neutralizing, surface-reacting, and immune-precipitating monoclonal antibodies. The predicted perturbations in the secondary structure introduced by the amino acid substitutions correlate with the extent of loss of reactivity with monoclonal antibodies in various immunoassays. 相似文献
2.
Antigenic analysis of a major neutralization site of herpes simplex virus glycoprotein D, using deletion mutants and monoclonal antibody-resistant mutants. 总被引:15,自引:14,他引:1
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M I Muggeridge V J Isola R A Byrn T J Tucker A C Minson J C Glorioso G H Cohen R J Eisenberg 《Journal of virology》1988,62(9):3274-3280
Herpes simplex virus glycoprotein D is a component of the virion envelope and appears to be involved in attachment, penetration, and cell fusion. Monoclonal antibodies against this protein can be arranged in groups on the basis of a number of biological and biochemical properties. Group I antibodies are type common, have high complement-independent neutralization titers, and recognize discontinuous (conformational) epitopes; they are currently being used in several laboratories to study the functions of glycoprotein D. We have used a panel of neutralization-resistant mutants to examine the relationships between these antibodies in detail. We found that they can be divided into two subgroups, Ia and Ib, such that mutations selected with Ia antibodies have little or no effect on binding and neutralization by Ib antibodies and vice versa. In addition, Ia antibodies are able to bind deletion and truncation mutants of glycoprotein D that Ib antibodies do not recognize, suggesting that their epitopes are physically distinct. However, with one exception, Ia and Ib antibodies block each other strongly in binding assays with purified glycoprotein D, whereas antibodies from other groups have no effect. We have therefore defined the sum of the Ia and Ib epitopes as antigenic site 1. 相似文献
3.
Characterization of a herpes simplex virus type 2 75,000-molecular-weight glycoprotein antigenically related to herpes simplex virus type 1 glycoprotein C. 总被引:22,自引:17,他引:5
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Evidence is presented that the herpes simplex virus type 2 glycoprotein previously designated gF is antigenically related to herpes simplex virus type 1 gC (gC-1). An antiserum prepared against type 1 virion envelope proteins immunoprecipitated gF of type 2 (gF-2), and competition experiments revealed that the anti-gC-1 component of the antiserum was responsible for the anti-gF-2 cross-reactivity. An antiserum prepared against fully denatured purified gF-2, however, and three anti-gF-2 monoclonal antibodies failed to precipitate any type 1 antigen, indicating that the extent of cross-reactivity between gC-1 and gF-2 may be limited. Several aspects of gF-2 synthesis and processing were investigated. Use of the enzymes endo-beta-N-acetylglucosaminidase H and alpha-D-N-acetylgalactosaminyl oligosaccharidase revealed that the fully processed form of gF-2 (about 75,000 [75K] apparent molecular weight) had both complex-type N-linked and O-linked oligosaccharides, whereas newly synthesized forms (67K and 69K) had only high-mannose N-linked oligosaccharides. These last two forms were both reduced in size to 54K by treatment with endo-beta-N-acetylglucosaminidase H and therefore appear to differ only in the number of N-linked chains. Neutralization tests and radioiodination experiments revealed that gF-2 is exposed on the surfaces of virions and that the 75K form of gF-2 is exposed on cell surfaces. The similarities and differences of gF-2 and gC-1 are discussed in light of recent mapping results which suggest collinearity of their respective genes. 相似文献
4.
Characterization of baculovirus-expressed herpes simplex virus type 1 glycoprotein K. 总被引:2,自引:2,他引:0
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The DNA region encoding the complete herpes simplex virus type 1 (HSV-1) glycoprotein K (gK) was inserted into a baculovirus transfer vector, and recombinant viruses expressing gK were isolated. Four gK-related recombinant baculovirus-expressed peptides of 29, 35, 38, and 40 kDa were detected with polyclonal antibody to gK. The 35-, 38-, and 40-kDa species were susceptible to tunicamycin treatment, suggesting that they were glycosylated. The 38- and 40-kDa species corresponded to partially glycosylated precursor gK (pgK) and mature gK, respectively. The 29-kDa peptide probably represented a cleaved, unglycosylated peptide. The 35-kDa peptide probably represented a cleaved, glycosylated peptide that may be a precursor to pgK. Indirect immunofluorescence with polyclonal antibody to gK peptides indicated that the recombinant baculovirus-expressed gK was abundant on the surface of the insect cells in which it was expressed. Mice vaccinated with the baculovirus-expressed gK produced very low levels (< 1:10) of HSV-1 neutralizing antibody. Nonetheless, these mice were partially protected from lethal challenge with HSV-1 (75% survival). This protection was significant (P = 0.02). Despite some protection against death, gK-vaccinated mice showed no protection against the establishment of latency. Surprisingly, gK-vaccinated mice that were challenged ocularly with a stromal disease-producing strain of HSV-1 had significantly higher levels of ocular disease (herpes stromal keratitis) than did mock-vaccinated mice. In summary, this is the first report to show that vaccination with HSV-1 gK can provide protection against lethal HSV-1 challenge and that vaccination with an HSV-1 glycoprotein can significantly increase the severity of HSV-1-induced ocular disease. 相似文献
5.
Genetic analysis of type-specific antigenic determinants of herpes simplex virus glycoprotein C.
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Herpes simplex virus type 1 (HSV-1) glycoprotein C (gC-1) elicits a largely serotype-specific immune response directed against previously described determinants designated antigenic sites I and II. To more precisely define these two immunodominant antigenic regions of gC-1 and to determine whether the homologous HSV-2 glycoprotein (gC-2) has similarly situated antigenic determinants, viral recombinants containing gC chimeric genes which join site I and site II of the two serotypes were constructed. The antigenic structure of the hybrid proteins encoded by these chimeric genes was studied by using gC-1- and gC-2-specific monoclonal antibodies (MAbs) in radioimmunoprecipitation, neutralization, and flow cytometry assays. The results of these analyses showed that the reactivity patterns of the MAbs were consistent among the three assays, and on this basis, they could be categorized as recognizing type-specific epitopes within the C-terminal or N-terminal half of gC-1 or gC-2. All MAbs were able to bind to only one or the other of the two hybrid proteins, demonstrating that gC-2, like gC-1, contains at least two antigenic sites located in the two halves of the molecule and that the structures of the antigenic sites in both molecules are independent and rely on limited type-specific regions of the molecule to maintain epitope structure. To fine map amino acid residues which are recognized by site I type-specific MAbs, point mutations were introduced into site I of the gC-1 or gC-2 gene, which resulted in recombinant mutant glycoproteins containing one or several residues from the heterotypic serotype in an otherwise homotypic site I background. The recognition patterns of the MAbs for these mutant molecules demonstrated that (i) single amino acids are responsible for the type-specific nature of individual epitopes and (ii) epitopes are localized to regions of the molecule which contain both shared and unshared amino acids. Taken together, the data described herein established the existence of at least two distinct and structurally independent antigenic sites in gC-1 and gC-2 and identified subtle amino acid sequence differences which contribute to type specificity in antigenic site I of gC. 相似文献
6.
Extensive homology between the herpes simplex virus type 2 glycoprotein F gene and the herpes simplex virus type 1 glycoprotein C gene. 总被引:14,自引:11,他引:3
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The region of the herpes simplex virus type 2 (HSV-2) genome which maps colinearly with the HSV-1 glycoprotein C (gC) gene has been cloned, and the DNA sequence of a 2.29-kilobase region has been determined. Contained within this sequence is a major open reading frame of 479 amino acids. The carboxyterminal three-fourths of the derived HSV-2 protein sequence showed a high degree of sequence homology to the HSV-1 gC amino acid sequence reported by Frink et al. (J. Virol. 45:634-647, 1983). The amino-terminal region of the HSV-2 sequence, however, showed very little sequence homology to HSV-1 gC. In addition, the HSV-1 gC sequence contained 27 amino acids in the amino-terminal region which were missing from the HSV-2 protein. Computer-assisted analysis of the hydrophilic and hydrophobic properties of the derived HSV-2 sequence demonstrated that the protein contained structures characteristic of membrane-bound glycoproteins, including an amino-terminal signal sequence and carboxy-terminal hydrophobic transmembrane domain and charged cytoplasmic anchor. The HSV-2 protein sequence also contained seven putative N-linked glycosylation sites. These data, in conjunction with mapping studies of Para et al. (J. Virol. 45:1223-1227, 1983) and Zezulak and Spear (J. Virol. 49:741-747, 1984), suggest that the protein sequence derived from the HSV-2 genome corresponds to gF, the HSV-2 homolog of HSV-1 gC. 相似文献
7.
8.
Cysteine mutants of herpes simplex virus type 1 glycoprotein D exhibit temperature-sensitive properties in structure and function. 总被引:6,自引:5,他引:1
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We previously constructed seven mutations in the gene for glycoprotein D (gD) of herpes simplex virus type 1 in which the codon for one of the cysteine residues was replaced by a serine codon. Each of the mutant genes was cloned into a eucaryotic expression vector, and the proteins were transiently expressed in mammalian cells. We found that alteration of any of the first six cysteine residues had profound effects on protein conformation and oligosaccharide processing. In this report, we show that five of the mutant proteins exhibit temperature-sensitive differences in such properties as aggregation, antigenic conformation, oligosaccharide processing, and transport to the cell surface. Using a complementation assay, we have now assessed the ability of the mutant proteins to function in virus infection. This assay tests the ability of the mutant proteins expressed from transfected plasmids to rescue production of infectious virions of a gD-minus virus, F-gD beta, in Vero cells. Two mutant proteins, Cys-2 (Cys-106 to Ser) and Cys-4 (Cys-127 to Ser), were able to complement F-gD beta at 31.5 degrees C but not at 37 degrees C. The rescued viruses, designated F-gD beta(Cys-2) and F-gD beta(Cys-4), were neutralized as efficiently as wild-type virus by anti-gD monoclonal antibodies, indicating that gD was present in the virion envelope in a functional form. Both F-gD beta(Cys-2) and F-gD beta(Cys-4) functioned normally in a penetration assay. However, the infectivity of these viruses was markedly reduced compared with that of the wild type when they were preincubated at temperatures above 37 degrees C. The results suggest that mutations involving Cys-106 or Cys-127 in gD-1 confer a temperature-sensitive phenotype on herpes simplex virus. These and other properties of the cysteine-to-serine mutants allowed us to predict a disulfide bonding pattern for gD. 相似文献
9.
The herpes simplex virus type I DNA polymerase. Polypeptide structure and antigenic domains 总被引:7,自引:0,他引:7
Polyclonal antibodies responding specifically to the N-terminal, central and C-terminal polypeptide domains of the herpes simplex virus type I (HSV-1) DNA polymerase of strain Angelotti were generated. Each of the five different rabbit antisera reacted specifically with a viral 132 +/- 5-kDa polypeptide as shown by immunoblot analysis. Enzyme binding and inhibition studies revealed that antibodies raised to the central and the C-terminal domains of the protein inhibited the polymerizing activity by 70-90%, respectively, which is well in line with the proposed site of the catalytic center of the enzyme and with the possible involvement of these polypeptide chains in DNA-protein interactions. In agreement with this, antibodies directed towards the N-terminal domain bound to the enzyme without effecting the enzymatic activity. The strong binding but low inhibitory properties of antibodies directed to the polypeptide region between residues 1072 and 1146 confirms previous suggestions that these C-terminal sequences, which share no homology to the Epstein-Barr virus DNA polymerase, are less likely involved in the building of the polymerase catalytic site. Antibodies, raised to the very C terminus of the polymerase (EX3), were successfully used to identify a single 132 +/- 5-kDa polypeptide, which coeluted with the HSV DNA polymerase activity during DEAE-cellulose chromatography, and were further shown to precipitate a major viral polypeptide of identical size. From the presented data it can be concluded that the native enzyme consists of a single polypeptide with a size predicted from the long open reading frame of the HSV-1 DNA polymerase gene. 相似文献
10.
M P Williamson B K Handa M J Hall 《International journal of peptide and protein research》1986,27(5):562-568
The peptide 6-amino caproyl-Pro-Ser-Leu-Lys-Met-Ala-Asp-Pro-Asn-Arg-Phe-Arg-Gly-Lys-Asp-Leu- Pro-6- amino caproate has been synthesized and its secondary structure has been investigated by 1H n.m.r. at 400 MHz. Resonances were assigned from 2D NOESY and COSY spectra, and the secondary structure was determined using NOEs, three-bond coupling constants, and exchange rates of amide protons. The peptide has two tight turns centered on the Pro-Asn and Arg-Gly pairs. The relationship between the secondary structure found here and the antigenic nature of the peptide is discussed. 相似文献
11.
Expression of herpes simplex virus type 1 glycoprotein D deletion mutants in mammalian cells. 总被引:21,自引:15,他引:6
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G H Cohen W C Wilcox D L Sodora D Long J Z Levin R J Eisenberg 《Journal of virology》1988,62(6):1932-1940
Glycoprotein D (gD) is a viron envelope component of herpes simplex virus types 1 and 2. We have previously defined seven monoclonal antibody (MAb) groups which recognize distinct epitopes on the mature gD-1 protein of 369 amino acids. MAb groups VII, II, and V recognize continuous epitopes at residues 11-19, 272-279, and 340-356, respectively. MAb groups I, III, IV, and VI recognize discontinuous epitopes. Recent studies have focused on epitopes I, III, and VI. Using truncated forms of gD generated by recombinant DNA methods and proteolysis, epitopes III, IV, and VI were located within amino acids 1-233. A portion of discontinuous epitope I was located in a region within residues 233-275. For this study, we used recombinant DNA methods to create mutations in the gD-1 gene and studied the effects of those mutations on gD as expressed in mammalian cells. Plasmid pRE4, containing the coding sequence of gD-1 and the Rous sarcoma virus long terminal repeat promoter, was transfected into mammalian cells. The expressed protein, gD-1-(pRE4), was identical in size and antigenic properties to gD-1 from infected cells. Six in-frame deletion mutations were subsequently constructed by using restriction enzymes to excise portions of the gD-1 gene. Plasmids carrying these mutated forms were transfected into cells, and the corresponding proteins were examined at 48 h posttransfection for antigenicity and glycosylation patterns. Three deletions of varying size were located downstream of residue 233. Analysis of these mutants showed that amino acids within the region 234-244 were critical for binding of DL11 (group I), but not for other MAb groups. Three other deletion mutants lost all ability to bind MAbs which recognize discontinuous epitopes. In addition, much of the gD expressed by these mutants was observed to migrate as high-molecular-weight aggregated forms in nondenaturing gels. Each of these mutations involved the loss of a cysteine residue, suggesting that disulfide linkages play an essential role in the formation of discontinuous epitopes. The extent of glycosylation of the mutant gD molecules accumulated at 48 h posttransfection suggested altered carbohydrate processing. In one case, there was evidence for increased O-linked glycosylation. Those proteins which had lost a cysteine residue as part of the deletion did not accumulate molecules processed beyond the high-mannose stage. The results suggest that carbohydrate processing during synthesis of gD is very sensitive to alterations in structure, particularly changes involving cysteine residues. 相似文献
12.
Detailed analysis of the portion of the herpes simplex virus type 1 genome encoding glycoprotein C. 总被引:81,自引:69,他引:12
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We previously showed that the right third of HindIII fragment L (0.59 to 0.65) of herpes simplex virus type 1 (HSV-1) encodes a family of mRNAs some members of which appear to be related by splicing. In the experiments described in this communication, we determined the nucleotide sequence of the DNA encoding this mRNA family and precisely located the mRNAs associated with this DNA sequence. The major mRNA species is unspliced and encoded by a 2.520-nucleotide region. Just upstream of the 5' end are TATA and CAT box sequences characteristic of HSV-1 promoters. The 3' end maps near a region containing a nominal polyadenylation signal. Three minor species (2,400, 2,200, and 1,900 bases, respectively) appear to share a very short leader sequence with the 5' end of the major mRNA and are then encoded by uninterrupted DNA sequences beginning about 100, 400, and 625 bases downstream of the 5' end of the major unspliced mRNA. These positions map at or very near positions which agree reasonably well with consensus splice acceptor sequences. The fourth mRNA is encoded by a contiguous 730-nucleotide sequence at the 3' end of the major unspliced mRNA and has its 5' end just downstream of recognizable TATA and CAT box sequences. We suggest that this mRNA is controlled by its own promoter. The nucleotide sequence data, in combination with the mRNA localization, demonstrate four potential polypeptides encoded by the region. The largest is 1,569 bases long and defines a 523-amino acid protein with sequence features characteristic of a glycoprotein. This was confirmed to be HSV-1 glycoprotein C by immune precipitation of the in vitro translation product of the major unspliced mRNA, performed with a polyspecific antibody to HSV-1 envelope glycoproteins (anti-env-1 serum), and by comparison of tryptic peptides of this translation product with those of authentic HSV-1 glycoprotein C. Polypeptides encoded by some of the minor species also were tentatively identified. 相似文献
13.
Pathogenicity in mice of herpes simplex virus type 2 mutants unable to express glycoprotein C. 总被引:9,自引:8,他引:1
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Herpes simplex virus type 2 (HSV-2) mutants that were unable to express glycoprotein C (gC-2) were isolated. Deletions were made in a cloned copy of the gC-2 gene, and recombinant viruses containing these deletions were screened by using an immunoreactive plaque selection protocol. The viruses did not display a syncytial phenotype. Intravaginal inoculation of BALB/cJ mice with one of the HSV-2 gC-2- viruses produced local inflammation followed by a lethal spread of the viral infection into the nervous system in a manner identical to that produced by parental HSV-2 strain 333. Similarly, intracerebral inoculation of DBA-2 mice with the gC-2- virus produced a lethal neurological disease paralleling that caused by HSV-2 strain 333. These results indicate that gC-2 is not required for the spread of HSV-2 infections in mice. 相似文献
14.
Molecular basis of the glycoprotein C-negative phenotypes of herpes simplex virus type 1 mutants selected with a virus-neutralizing monoclonal antibody. 总被引:12,自引:10,他引:2
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Previously (Holland et al., J. Virol. 52:566-574, 1984; Kikuchi et al., J. Virol. 52:806-815, 1984) we described the isolation and partial characterization of over 100 herpes simplex virus type 1 mutants which were resistant to neutralization by a pool of glycoprotein C- (gC) specific monoclonal antibodies. The genetic basis for the inability of several of these gC- mutants to express an immunoreactive envelope form of gC is reported here. Comparative nucleotide sequence analysis of the gC gene of the six mutants gC-3, gC-8, gC-49, gC-53, gC-85, and synLD70, which secrete truncated gC polypeptides, with that of the wild-type KOS 321 gC gene revealed that these mutant phenotypes were caused by frameshift or nonsense mutations, resulting in premature termination of gC translation. Secretion of the gC polypeptide from cells infected with these mutants was due to the lack of a functional transmembrane anchor sequence. The six secretor mutants were tested for suppression of amber mutations in mixed infection with a simian virus 40 amber suppressor vector. Mutant gC-85 was suppressed and produced a wild-type-sized membrane-bound gC. Nucleotide sequence analysis of the six gC deletion mutants gC-5, gC-13, gC-21, gC-39, gC-46, and gC-98 revealed that they carried identical deletions which removed 1,702 base pairs of the gC gene. The deletion, which was internal to the gC gene, removed the entire gC coding sequence and accounted for the novel 1.1-kilobase mRNA previously seen in infections with these mutants. The mutant gC-44 was previously shown to produce a membrane-bound gC protein indistinguishable in molecular weight from wild-type gC. This mutant differed from wild-type virus in that it had reduced reactivity with virus-neutralizing monoclonal antibodies. Nucleotide sequence analysis of the gC gene of mutant gC-44 demonstrated a point mutation which changed amino acid 329 of gC from a serine to a phenylalanine. 相似文献
15.
16.
Varicella-zoster virus complements herpes simplex virus type 1 temperature-sensitive mutants. 总被引:17,自引:13,他引:4
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Varicella-zoster virus (VZV) can complement temperature-sensitive mutants of herpes simplex virus. Of seven mutants tested, two, carrying mutations in the immediate-early ICP4 and ICP27 proteins, were complemented. This complementation was not seen in coinfections with adenovirus type 5 or cytomegalovirus. Following transfection into CV-1 cells, a DNA fragment containing the VZV short repeat sequence complemented the ICP4 mutant. These data demonstrate a functional relationship between VZV and herpes simplex virus and have allowed localization of a putative VZV immediate-early gene. 相似文献
17.
Passive immune protection by herpes simplex virus-specific monoclonal antibodies and monoclonal antibody-resistant mutants altered in pathogenicity. 总被引:3,自引:11,他引:3
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Virus-neutralizing monoclonal antibodies specific for 13 different genetically defined epitopes of glycoproteins gC, gB, and gD of herpes simplex virus type 1, strain KOS-321, were compared for their ability to provide passive immunity to DBA-2 mice challenged intracranially. Protection was highly specific, since individual monoclonal antibodies failed to protect against infection with monoclonal antibody-resistant (mar) mutants altered in the single epitope recognized by the injected antibody. The dose-response kinetics of passive immunity paralleled the in vitro neutralization titers for each antibody. No correlation was observed between immune protection and antibody isotype or complement-dependent in vitro neutralization titers. This suggests that virus neutralization was not the protective mechanism. In general, antibodies reactive with epitopes of gC were protective at the lowest antibody doses, antibodies specific for gB were less efficient in providing immunity, and antibodies against gD were the least effective. mar mutants with single epitope changes in gC and multiple epitope changes in gB showed highly reduced pathogenicity, requiring up to 5 X 10(6) PFU to kill 50% of infected animals. These findings indicated that antigenic variation affects virus growth and spread in the central nervous system. Thus, mutations which affect antigenic structure also can alter virus pathogenicity. The alteration of these epitopes does not, however, appreciably reduce the development of resistance to infection. Infection of mice with these mutants or inoculation of mice with UV-inactivated, mutant-infected cells before challenge rendered the animals resistant to infection with wild-type herpes simplex virus type 1. 相似文献
18.
DNA synthesis and DNA polymerase activity of herpes simplex virus type 1 temperature-sensitive mutants. 总被引:43,自引:33,他引:10
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Fifteen temperature-sensitive mutants of herpes simplex virus type 1 were studied with regard to the relationship between their ability to synthesize viral DNA and to induce viral DNA polymerase (DP) activity at permissive (34 C) and nonpermissive (39 C) temperatures. At 34 C, all mutants synthesized viral DNA, while at 39 C four mutants demonstrated a DNA+ phenotype, three were DNA+/-, and eight were DNA-. DNA+ mutants induced levels of DP activity similar to thhose of the wild-type virus at both temperatures, and DNA+/- mutants induced reduced levels of DP activity at 39 C but not at 34 C. Among the DNA- mutants three were DP+, two were DP+/-, and three showed reduced DP activity at 34 C with no DP activity at 39 C. DNA-, DP- mutants induced the synthesis of a temperature-sensitive DP as determined by in vivo studies. 相似文献
19.
Fine structure analysis of type-specific and type-common antigenic sites of herpes simplex virus glycoprotein D 总被引:5,自引:15,他引:5
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B Dietzschold R J Eisenberg M Ponce de Leon E Golub F Hudecz A Varrichio G H Cohen 《Journal of virology》1984,52(2):431-435
The fine structure of the antigenic determinants of herpes simplex virus type 1 and 2 glycoprotein D (gD) was analyzed to determine whether structural differences underlie the differential immunogenicity of these glycoproteins. A region common to herpes simplex virus type 1 and 2 gD (amino acid residues 11 to 19) and two sites specific for herpes simplex virus type 2 gD (one determined by proline at position 7, the other determined by asparagine at position 21) were localized within the N-terminal 23 amino acids of gD by synthesis of peptides and comparison of their cross-reactivity with antisera raised to herpes simplex virus type 1 and 2 gD. The secondary structure of these peptides, as predicted by computer analysis, is discussed in relation to their immunogenicity. 相似文献
20.
利用PCR拼接技术,合成含单纯疱疹病毒II型糖蛋白G(gG2)抗原表位(氨基酸序列第561~578位)的片段,并进一步利用基因工程技术获得该表位的双拷贝片段,克隆入pET-KDO表达载体进行原核表达。经IPTG诱导后,高效表达出分子量大小约为39,000D的融合蛋白,经Western-blot检测具有良好的抗原性。表达的融合蛋白经凝血酶切割和亲和层析纯化,得到双拷贝gG2(561~578aa)目的蛋白,经ELISA检测具有良好的灵敏度和特异性。该重组抗原的构建和表达可用于HSV-2特异性血清学诊断的研究。 相似文献