Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

A library of pseudorabies virus (PRV) DNA fragments was constructed in the expression cloning vector lambda gt11. The library was screened with antisera which reacted with mixtures of PRV proteins to isolate recombinant bacteriophages expressing PRV proteins. By the nature of the lambda gt11 vector, the cloned proteins were expressed in Escherichia coli as beta-galactosidase fusion proteins. The fusion proteins from 35 of these phages were purified and injected into mice to raise antisera. The antisera were screened by several different assays, including immunoprecipitation of [14C]glucosamine-labeled PRV proteins. This method identified phages expressing three different PRV glycoproteins: the secreted glycoprotein, gX; gI; and a glycoprotein that had not been previously identified, which we designate gp63. The gp63 and gI genes map adjacent to each other in the small unique region of the PRV genome. The DNA sequence was determined for the region of the genome encoding gp63 and gI. It was found that gp63 has a region of homology with a herpes simplex virus type 1 (HSV-1) protein, encoded by US7, and also with varicella-zoster virus (VZV) gpIV. The gI protein sequence has a region of homology with HSV-1 gE and VZV gpI. It is concluded that PRV, HSV, and VZV all have a cluster of homologous glycoprotein genes in the small unique components of their genomes and that the organization of these genes is conserved.
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PMID:Use of lambda gt11 to isolate genes for two pseudorabies virus glycoproteins with homology to herpes simplex virus and varicella-zoster virus glycoproteins. 301 84

To localize the genes for the major glycoproteins of equine herpesvirus 1 (EHV-1), a library of the EHV-1 genome was constructed in the lambda gt11 expression vector. Recombinant bacteriophage expressing EHV-1 glycoprotein epitopes as fusion products with beta-galactosidase were detected by immunoscreening with monoclonal antibodies specific for each of six EHV-1 glycoproteins. Seventy-four recombinant lambda gt11 clones reactive with EHV-1 monoclonal antibodies were detected among 4 X 10(5) phage screened. Phage expressing determinants on each of the six EHV-1 glycoproteins were represented in the library. Herpesviral DNA sequences contained in lambda gt11 recombinants expressing epitopes of EHV-1 glycoproteins were used as hybridization probes for mapping insert sequences on the viral genome. Genes for five EHV-1 glycoproteins (gp2, gp10, gp13, gp14, and gp21/22a) mapped to the genome L component; only one EHV-1 glycoprotein (gp17/18) was expressed from the unique S region of the genome where genes of several major glycoproteins of other herpesviruses have been located. Two glycoproteins of EHV-1, gp13 and gp14, mapped to positions colinear with genes of major glycoproteins identified in several other alphaherpesviruses (gC- and gB-like glycoproteins, respectively). The genomic locations of other EHV-1 glycoproteins indicated the existence of major glycoproteins of EHV-1 (gp2, gp10, and gp21/22a) for which no genetic homologs have yet been detected in other herpesviruses. The results confirm the general utility of the lambda gt11 expression system for localizing herpesvirus genes and suggest that the genomic positioning of several high-abundance glycoproteins of EHV-1 may be different from that of the prototype alphaherpesvirus, herpes simplex virus.
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PMID:Use of lambda gt11 and monoclonal antibodies to map the genes for the six major glycoproteins of equine herpesvirus 1. 303 8

We have looked for conserved DNA sequences between four herpes simplex virus type 1 (HSV-1) glycoprotein genes encoding gB, gC, gD, and gE and pseudorabies virus (PRV) DNA, HSV-1 DNA fragments representing these four glycoprotein-coding sequences were hybridized to restriction enzyme fragments of PRV DNA by the Southern blot procedure. Specific hybridization was observed only when HSV-1 gB DNA was used as probe. This region of hybridization was localized to a 5.2-kilobase (kb) region mapping at approximately 0.15 map units on the PRV genome. Northern blot (RNA blot) analysis, with a 1.2-kb probe derived from this segment, revealed a predominant hybridizing RNA species of approximately 3 kb in PRV-infected PK15 cells. DNA sequence analysis of the region corresponding to this RNA revealed a single large open reading frame with significant nucleotide homology with the gB gene of HSV-1 KOS 321. In addition, the beginning of the sequenced PRV region also contained the end of an open reading frame with amino acid homology to HSV-1 ICP 18.5, a protein that may be involved in viral glycoprotein transport. This sequence partially overlaps the PRV gB homolog coding sequence. We have shown that the PRV gene with homology to HSV-1 gB encoded the gII glycoprotein gene by expressing a 765-base-pair segment of the PRV open reading frame in Escherichia coli as a protein fused to beta-galactosidase. Antiserum, raised in rabbits, against this fusion protein immunoprecipitated a specific family of PRV glycoproteins of apparent molecular mass 110, 68, and 55 kilodaltons that have been identified as the gII family of glycoproteins. Analysis of the predicted amino acid sequence indicated that the PRV gII protein shares 50% amino acid homology with the aligned HSV-1 gB protein. All 10 cysteine residues located outside of the signal sequence, as well as 4 of 6 potential N-linked glycosylation sites, were conserved between the two proteins. The primary protein sequence for HSV-1 gB regions known to be involved in the rate of virus entry into the cells and cell-cell fusion, as well as regions known to be associated with monoclonal antibody resistance, were highly homologous with the PRV protein sequence. Furthermore, monospecific antibody made against PRV gII immunoprecipitated HSV-1 gB from infected cells. Taken together, these findings suggest significant conservation of structure and function between the two proteins and may indicate a common evolutionary history.
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PMID:The pseudorabies virus gII gene is closely related to the gB glycoprotein gene of herpes simplex virus. 303 63

gG.2 glycoprotein was purified by H966 monoclonal antibodies linked to Sepharose from herpes simplex virus type 2-infected HEp-2 cells labeled with [3H] glucosamine. The glycoprotein was subjected to Pronase digestion and the glycopeptides were fractionated by Con A-Sepharose in a major fraction (88.5% of total radioactivity) unbound to the lectin gel and in a minor species which bound to the lectin as a N-linked diantennary oligosaccharide. Mild and strong acid hydrolysis of Con A-unbound and Con A-bound fractions revealed that (i) both species were highly sialylated; (ii) the Con A-unbound fraction contained mainly labeled N-acetylgalactosamine, as is the case for O-linked oligosaccharides; and (iii) the Con A-bound fraction carried the vast majority of the labeled N-acetylglucosamine present in gG.2. Three size classes of oligosaccharides were separated from mild alkaline borohydride-treated Con A-unbound glycopeptides, which accounted for about 80% of the radioactivity present in the fraction. Galactosaminitol was recovered as the major labeled product in the strong acid hydrolyzates of the oligosaccharides generated by reductive beta-elimination, indicating that they were O-glycosidically linked to the peptide backbone. Thin-layer and DEAE-Sephacel chromatography of the three O-linked oligosaccharide species indicated that disialylated tetrasaccharides and monosialylated trisaccharides were the major components, whereas neutral disaccharide was a minor component. Digestion with neuraminidase and beta-galactosidase of the O-linked oligosaccharides supported the idea that the common disaccharide core was mainly of the structure beta-galactosyl-N-acetylgalactosamine. The large occurrence of O-linked oligosaccharides differentiates this type 2-specific herpes simplex virus glycoprotein from the type-common herpesvirus glycoproteins gB, gC, and gD.
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PMID:Oligosaccharide chains of herpes simplex virus type 2 glycoprotein gG.2. 402 10

A plasmid-borne Herpes simplex virus type 1 (HSV-1) thymidine kinase (TK) gene (tk) was expressed in Escherichia coli by inserting a 203-bp lacL8/UV5 promoter-operator segment, in frame, 53 bp 5' to the native tk translational start codon. The hybrid gene created by this fusion encodes a polypeptide which has 25 additional amino acids on the amino terminus of the HSV-1 TK protein and phenotypically complements a tdk- mutation of E. coli. This fusion polypeptide has been characterized by maxicell, immunoprecipitation, and native gel techniques, and its activity is inhibited by anti-HSV-1 antibody. In a tk expressor strain containing a F' lacIq (which overproduces the lactose repressor), the isopropyl-beta-D-thiogalactoside (IPTG) causes greater than 1000-fold coordinate induction of the plasmid-encoded TK and chromosomal beta-galactosidase activities. Pulse-chase induction demonstrates the fused TK polypeptide to be as stable as beta-galactosidase. HSV-1 tk-specific RNA isolated from this bacterial strain has a short half-life characteristic of bacterial messages.
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PMID:Regulated high-level expression of the Herpes simplex type I thymidine kinase gene in Escherichia coli. 608 61

We have taken a new approach to identify and fine map previously undescribed herpes simplex virus (HSV) functions. In experiments described in this report the antibiotic coumermycin A1 was used to select two HSV type 1 BamHI fragments cloned in pBR322 that confer partial resistance to drug-susceptible Escherichia coli. The genes encoding these HSV functions have been designated cour-1 and cour-2 and have been fine mapped to the HSV sequences. HSV-cour1 is located at the left end of BamHI-F near HSV type 1 genomic map coordinate 0.645. cour-2 maps to BamHI-M', which is a 159-base-pair internal component of the alpha ICP4-coding sequence located in the reiterated sequences of the S component. In pBR322 both inserts apparently rely on the tet promoter for expression. Additionally, cour-2 functions when present as a BamHI insert in pUC7. The analysis of cour-2 "maxi" cell proteins reveals the presence of proteins produced by the fusion of HSV-1 BamHI-M' sequences and the sequences of the vector genes, i.e., the major tet product for pBR322 and the modified beta-galactosidase for pUC7. These data suggest that the development of bacterial assays for fusion products of eucaryotic DNA open reading frames in plasmid vectors may be a useful technique for initiating gene function studies.
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PMID:Herpes simplex virus cloned DNA fragments induce coumermycin A1 resistance in Escherichia coli. 609 66

DNA fragments encoding structural information of the Herpes simplex virus type 1 (HSV-1) glycoprotein C (gC) gene were cloned into pUC plasmids [Vieira and Messing, Gene 19 (1982) 259-268]. None of the hybrid plasmids were able to direct the synthesis of significant amounts of gC related peptides. Several of the plasmid-bearing strains, however, exhibited inhibition characteristics which can be correlated with the presence on the plasmid of specific gC gene sequences. After insertion of gC DNA fragments into expression vector pMF2 between phage lambda repressor gene cI and lacZ, significant amounts of cI::gC::beta-galactosidase fusion proteins are synthesized. These tripartite fusion proteins are immunologically reactive with anti-HSV-1 antisera. The expression system based on pMF2 can be generally used to identify and express foreign antigens in Escherichia coli.
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PMID:Expression of Herpes simplex virus type 1 glycoprotein C antigens in Escherichia coli. 609 9

Herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) cause both persistent and latent infections, including recurrent cutaneous disease, lethal neonatal disease, central nervous system disease and other clinical syndromes. Modified live vaccines or conventionally prepared subunit vaccines have generally been unsuccessful in the treatment of HSV-1 and HSV-2 infections from the standpoints of safety and efficacy. It has been established that HSV-1 and HSV-2 infectivity may be neutralized in vitro with antisera directed specifically against each of the four major glycoproteins of the virus (gA/gB, gC, gD and gE) and antisera against glycoprotein gD, of either HSV-1 or HSV-2, are capable of neutralizing both HSV-1 and HSV-2 infectivity in vitro and in vivo. We have previously reported on the identification, DNA sequence and expression at low level in Escherichia coli of the gD gene of HSV-1 strain Patton. Here we describe construction of a hybrid gene encoding a chimaeric protein containing HSV-1 gD, bacteriophage lambda Cro and E. coli beta-galactosidase (gD-beta-gal) protein, which is expressed at high level in E. coli. Moreover, the chimaeric protein elicits antibodies in rabbits that not only immunoprecipitate gD from cells infected with HSV-1 and HSV-2 but also neutralize HSV-1 and HSV-2 infectivity in vitro.
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PMID:An immunologically active chimaeric protein containing herpes simplex virus type 1 glycoprotein D. 629 36

We have used elements of the E. coli lactose (lac) operon to produce a collection of herpes simplex virus types 1 and 2 glycoprotein D (gD-1 and gD-2) antigens. Our approach employed recombinant DNA techniques to construct plasmids with various segments of the gD-1 and gD-2 coding sequences fused to the lacZ gene. Such hybrid genes were expressed in a regulated manner in E. coli by joining them to the lac promoter-operator region. Efficient translation of these hybrid genes was facilitated by incorporating a coding sequence specifying a short peptide leader (lambda cro) in the plasmid expression vectors resulting in synthesis of chimeric Cro-gD-beta-galactosidase proteins. In addition, insertion of synthetic translation terminators at the junction of gD and lacZ enabled us to produce specific truncated gD polypeptide sequences unfused to beta-galactosidase. The gD antigens produced in E. coli were not glycosylated and were generally recovered as dense insoluble aggregates. Proteins containing portions of gD-1 or gD-2 were analyzed by immunoprecipitation using anti-HSV rabbit serum and a number of monoclonal antibodies recognizing different epitopes of gD-1. Initial animal studies were done with antigens that reacted with neutralizing antisera or monoclonal antibodies. When these bacterially produced proteins were injected into rabbits, antibodies were produced that specifically immunoprecipitated authentic gD polypeptides and neutralized the infectivity of both virus types. These studies suggest that gene fusion techniques can be used to produce immunogenic proteins in large quantity. These polypeptides are not only useful in analyses of gene structure and function, but also can provide novel diagnostic reagents and well-defined pure antigens for vaccine development.
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PMID:Bacterial synthesis of herpes simplex virus types 1 and 2 glycoprotein D antigens. 633 Feb 15

Activity and intracellular distribution of lysosomal glucosidases (beta-glucosidase and beta-galactosidase) of the cornea as well as membrane permeability were studied in experimental herpetic keratitis. It was demonstrated that the action of herpes simplex entailed labilization of the lysosomal membranes of the cornea together with the decreased strength of lysosomal association with the enzymes thereby favouring the increased enzyme-substrate contact and excess destruction of cellular and exocellular glycosaminoglycans. The data obtained suggest that lysosomes participate in the pathogenesis of herpetic keratitis.
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PMID:[Corneal lysosomes in experimental ophthalmoherpes]. 678 7


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