Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.23 (beta-galactosidase)
 14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Herpes simplex virus (HSV) glycoprotein gD is a major component of the virion envelope and is thought to play an important role in the initial stages of viral infection and stimulates the production of high titers of neutralizing antibodies. We assumed that gD plays an essential role in virus replication, and so to complement viruses with mutations in the gD gene we constructed a cell line, denoted VD60, which is capable of expressing high levels of gD after infection with HSV. A recombinant virus, designated F-gD beta, in which sequences encoding gD and a nonessential glycoprotein, gI, were replaced by Escherichia coli beta-galactosidase sequences, was selected on the basis that it produced blue plaques on VD60 cell monolayers under agarose overlays containing 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside (X-Gal). F-gD beta was able to replicate normally on complementing VD60 cells. However, F-gD beta was unable to form plaques on noncomplementing Vero cells. Virions lacking gD were produced in normal amounts by Vero cells infected with F-gD beta, and the virus particles were distributed throughout the cytoplasm and on the cell surface, suggesting that gD is not essential for HSV envelopment and egress. Virions lacking gD were able to bind to cells, but were unable to initiate synthesis of viral early polypeptides. Plaque production of F-gD beta particles lacking gD was enhanced by polyethylene glycol treatment, suggesting that gD is essential for penetration of HSV into cells. Other HSV glycoproteins have been implicated in the entry of virus into cells, and thus this process appears to involve multiple interactions at the cell surface.
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PMID:A herpes simplex virus mutant in which glycoprotein D sequences are replaced by beta-galactosidase sequences binds to but is unable to penetrate into cells. 283 3

A defective herpes simplex virus 1 (HSV-1) vector, pHSVlac, has been developed that contains a transcription unit that places the Escherichia coli lacZ gene under the control of the HSV-1 immediate early 4/5 promoter. The vector pHSVlac was propagated with the HSV-1 temperature-sensitive mutant ts K as helper virus. Infection of neurons from rat superior cervical ganglia and dorsal root ganglia in primary culture resulted in stable expression of high levels of beta-galactosidase without cell death. These HSV-1 vectors should be useful for introducing genes into postmitotic cells, such as neurons, in vitro and in vivo.
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PMID:A defective HSV-1 vector expresses Escherichia coli beta-galactosidase in cultured peripheral neurons. 284 86

Purified preparations of herpes simplex virus type 1 Angelotti were digested with the exoglycosidases sialidase, beta-galactosidase, N-acetyl-beta-D-glucosaminidase and alpha-mannosidase, and with the endoglycosidases Endo-H and Endo-F. It was found that treatment of virions with Endo-F specifically decreased viral infectivity by a factor of 10. This reduction in titre was not associated with any measurable differences in virus adsorption, suggesting a role of N-linked complex type oligosaccharide chains in penetration. In contrast, a reduction in titre observed upon digestion of virions with exoglycosidases could be attributed to a proteolytic contamination in these enzyme preparations. Treatment of virions with Endo-H, demonstrated to be free of proteolytic contamination, did not reduce viral infectivity. Analysis of endoglycosidase-digested virions by monospecific antibodies and immunoblotting revealed a susceptibility of all four major glycoproteins (gC, gB, gE and gD) to Endo-F, but only gB was susceptible to Endo-H treatment. In contrast, of all the exoglycosidases used only sialidase was found to be active towards native viral glycoproteins. Upon analysis of endoglycosidase-digested virions we could not find any evidence for proteolysis, degradation or altered protein composition of viral envelopes. In contrast, vigorous inhibition of glycoprotein glycosylation by tunicamycin led to the formation of physically intact virions almost completely lacking all major glycoproteins. These data show that digestion of intact virions with glycosidases allows an analysis of the functional relevance of carbohydrate residues without any obvious alterations in the virion glycoprotein composition.
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PMID:Removal of N-linked carbohydrates decreases the infectivity of herpes simplex virus type 1. 284 61

We have inserted a modified Escherichia coli lacZ gene, placed under the control of herpes simplex virus alpha 4 or beta 8 regulatory signals, into the HSV-1 genome disrupting the viral thymidine kinase gene. Using beta-galactosidase as an in situ indicator of viral gene expression, we detected expression from these recombinant HSV in dermal and neural tissues of the BALB/c mouse. Our detection of beta-galactosidase expression in neuronal cells indicates that TK-deficient viruses are capable of invading mouse neuronal cells and expressing up to the beta class of gene product.
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PMID:Beta-galactosidase as a marker in the peripheral and neural tissues of the herpes simplex virus-infected mouse. 284 16

The expression of Herpes Simplex Virus 1 (HSV-1) glycoprotein C (gC), a well defined herpesvirus late gene, was studied by linking the promoter-regulatory region of this gene to the coding sequences for the bacterial enzyme, beta-galactosidase (beta-gal). A chimeric gene, containing the beta-gal gene under the control of gC sequences from -1350 to +30 relative to the mRNA start site, was inserted by homologous recombination into the thymidine kinase (TK) locus of the HSV-1 genome. Selection of the TK- recombinant virus by plaque assay was facilitated by addition of a beta-gal indicator to the agarose overlay. Recombinant virus containing the gC promoter-beta-gal chimeric gene faithfully expressed beta-gal as a viral late gene, as shown by the absence of beta-gal expression when viral DNA replication was inhibited with phosphonoacetic acid. In contrast, the inhibition of viral DNA replication had no effect on the expression of beta-gal when the beta-gal gene was under the control of the early HSV-1 TK promoter in a separate recombinant virus. Analysis of recombinant viruses containing 5' to 3' deletions in the gC regulatory region revealed no apparent difference in beta-gal expression as deletions extended from -1350 to -109 base-pairs (bp) before the RNA start site, demonstrating that sequences between -109 and +30 are sufficient for regulated gC expression in the viral genome. Analysis of the mRNA made by these recombinant viruses confirmed the results of the beta-gal assays, and demonstrated that the transcriptional start sites of the gC promoter-beta-gal chimeric genes were the same as the start site of the gC gene.
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PMID:The use of beta-galactosidase as a marker gene to define the regulatory sequences of the herpes simplex virus type 1 glycoprotein C gene in recombinant herpesviruses. 284 20

An in situ ELISA was performed directly on the adherent cell monolayer in order to determine the susceptibility of herpes simplex virus isolates to acyclovir. Various fixation procedures and antisera conjugated to different enzymes were tested. The use of glutaraldehyde for fixation and beta-galactosidase as a labelling enzyme was shown to give the best results. As with other currently used assays, 50% inhibitory doses were subject to an inoculum effect. The data obtained indicate that this assay is suitable for routine determination of herpes simplex virus susceptibility to antiviral drugs.
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PMID:Evaluation of herpes simplex virus susceptibility to acyclovir using an enzyme-linked immunosorbent assay. 284 62

The N-terminal fragment, comprising residues -5 to 55 of herpes simplex virus type 1 glycoprotein D was expressed as a beta-galactosidase fusion protein in Escherichia coli. This gD-fusion protein reacts with monoclonal antibody LP 14 directed against glycoprotein D of HSV. Antisera obtained after immunization of rabbits with purified gD-fusion protein react with HSV-1 gD in a Western blot and with N-terminal synthetic peptides of gD. In addition, these antisera are able to neutralize viral infectivity in vitro.
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PMID:Immunological properties of an N-terminal fragment of herpes simplex virus type 1 glycoprotein D expressed in Escherichia coli. 285 Jul 85

We have constructed a transient expression vector (pCAL) containing two reporter genes, chloramphenicol acetyltransferase (CAT) and beta-galactosidase (beta-gal) for use in studying herpes simplex virus type 1 (HSV-1) promoter activity in mammalian cells. The construct was designed to be useful in analyzing the simultaneous expression from two different promoters. To test the utility of the vector, we used three HSV-1 promoters that had been characterized previously by workers in this laboratory. Two are early (beta) promoters, for alkaline exonuclease and deoxyuridine triphosphate nucleotidohydrolase; the third promoter controls the major capsid protein transcript and is late (beta gamma). The two different kinetic classes of promoters were ligated in a divergent orientation into pCAL and transfected into rabbit skin fibroblast. Transfected cells were then superinfected with low multiplicities of HSV-1; 18 hr later, we observed the simultaneous expression of both marker genes under control of the respective promoters. The usefulness of such a transient expression reporter vector is discussed.
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PMID:A bi-functional reporter plasmid for the simultaneous transient expression assay of two herpes simplex virus promoters. 285 22

A series of plasmid vectors has been constructed for the regulated, high level expression of foreign genes in E. coli. The vectors express cloned genes under the control of the tac promoter, which is a hybrid of trp and lac promoter sequences. Some of our expression vectors carry in addition to the tac promoter, the efficient lacZ ribosome binding site followed by unique cloning sites. These vectors can be used to express cloned genes directly, i.e. in an unfused, mature form. A second type of vector provides, in addition to the above regulatory elements, a translation initiation codon (ATG) for the expression of genes which have been isolated in an incomplete form (for example: cDNA). A third type of vector allow readily the construction of gene fusions to the E. coli beta-galactosidase gene, which may stabilize otherwise unstable eukaryotic proteins, and thus allows the production of high amounts of specific antigens in E. coli. With the above vectors, several eukaryotic and viral proteins, including SV40 small tumor antigen, human fibroblast interferon and herpes simplex glycoproteins have been expressed.
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PMID:Plasmid vectors for the regulated, high level expression of eukaryotic genes in Escherichia coli. 298 46

We have fused immediate (alpha) and delayed (beta) early promoter-regulatory sequences taken from the cytomegalovirus (CMV) genome to Escherichia coli lacZ (beta-galactosidase) as an indicator gene to study regulated expression of these promoters. After transfection of human fibroblast cells with plasmid constructs carrying beta-galactosidase fusions, and subsequent infection with CMV, we have demonstrated that viral trans-acting functions up-regulate the expression of these genes in a temporally authentic manner. The alpha promoter is activated even when de novo protein synthesis is blocked and when UV-inactivated virus is used, suggesting that, as for herpes simplex virus type 1 (HSV-1), a virion structural protein is responsible for its up-regulation. We have found that HSV-1, as well as CMV, is capable of trans activating the CMV alpha promoter. The beta promoter is activated by CMV but is completely unresponsive to HSV-1 infection. The temporal synthesis of the alpha and beta promoters in the transient expression system conforms with their natural regulation during viral replication. The beta-galactosidase fusions we describe provide a most exquisitely sensitive indicator system for the study of cis- and trans-acting viral regulatory functions.
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PMID:Regulation of cytomegalovirus gene expression: alpha and beta promoters are trans activated by viral functions in permissive human fibroblasts. 299 44


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