Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A new series of double-selection plasmids containing recombinant genes expressing the neomycin phosphotransferase (NEO) of transposon Tn5 and mouse dihydrofolate reductase (DHFR) in mammalian cells is described. Activity of the recombinant DHFR gene varied more than 50-fold, depending on the location of the simian virus 40 72 base-pair repeat or enhancer, which is part of the promoter of the NEO unit. A NEO-DHFR module with the enhancer located at the 3' end of the DHFR gene was inserted into a plasmid containing four tandem head-to-tail copies of the hepatitis B virus (HBV) genome and the new plasmid was used to transform DHFR- Chinese hamster ovary cells. In one of the cell lines obtained, an unrearranged copy of the HBV tetramer could be amplified 300-fold by increasing selective pressure with methotrexate, resulting in a proportional increase of the synthesis of HBV surface antigen. Four different mRNAs detected in the amplified cell line probably encode HBV core protein, pre-S and surface antigens, and the X protein. As a result of the DNA amplification, synthesis of HBV proteins is no longer restricted to resting cells. Integrated plasmid sequences appear to be stable during the amplification process.
J Mol Biol 1986 Jun 05
PMID:A recombinant Chinese hamster ovary cell line containing a 300-fold amplified tetramer of the hepatitis B genome together with a double selection marker expresses high levels of viral protein. 302 26

The activity of the hepatitis B viral enhancer element was studied in various cell lines. This enhancer shows strict host and tissue specificity in that it is functional only in liver cells of human origin. Further, it requires trans-acting factor(s) present in liver cells for activity, and this activity is independent of hepatitis B virus gene products in the cell lines tested.
Mol Cell Biol 1986 Feb
PMID:The human hepatitis B virus enhancer requires trans-acting cellular factor(s) for activity. 302 64

Hepatitis B surface antigen (HBsAg), the major coat protein of hepatitis B virus, is also secreted from cells as a subviral particle, without concomitant cleavage of N-terminal amino acid sequences. We examined this unusual export process in a cell-free system and showed that the initial product of HBsAg biosynthesis is an integral transmembrane protein, with most or all of its C-terminal half on the lumenal side of the endoplasmic reticulum membrane. To study the nature of its topogenic signals, we synthesized fusion proteins between HBsAg and the nonsecreted protein alpha-globin. Fusion proteins in which approximately 100 amino acids of globin preceded all HBsAg sequences were successfully translocated in vitro; the same domain as in the wild-type HBsAg was transported into the vesicle lumen. Fusions in which the entire globin domain was C terminal were able to translocate both the C-terminal region of HBsAg and its attached globin domain. Thus, uncleaved signal sequences in p24s function to direct portions of the molecule across the membrane and are able to perform this function even when positioned in an internal protein domain.
Mol Cell Biol 1986 May
PMID:Hepatitis B surface antigen: an unusual secreted protein initially synthesized as a transmembrane polypeptide. 302 91

A novel expression system based on coinfection of cells with two recombinant vaccinia viruses has been developed. One recombinant vaccinia virus contained the bacteriophage T7 RNA polymerase gene under control of a vaccinia virus promoter. The second recombinant vaccinia virus contained a target gene of choice flanked by bacteriophage T7 promoter and termination sequences. Maximum expression of the target gene occurred when cells were infected with 10 PFU of each recombinant virus. Although T7 RNA polymerase synthesis began shortly after infection, the target gene was not expressed until late times and was largely inhibited when DNA replication was blocked. Target gene transcripts were analyzed by agarose gel electrophoresis and had the predicted size. With this system, Escherichia coli beta-galactosidase, hepatitis B virus surface antigen, and human immunodeficiency virus envelope proteins were made. In each case, the level of synthesis was greater than had previously been obtained with the more conventional recombinant vaccinia virus expression system.
Mol Cell Biol 1987 Jul
PMID:Use of a hybrid vaccinia virus-T7 RNA polymerase system for expression of target genes. 311 59

The S promoter, one of the major hepatitis B virus (HBV) promoters, directs the synthesis of mRNA for surface antigen. Transient expression studies revealed that this promoter is highly active in the Alexander hepatoma cell line but not in SK-Hep1 and HeLa cells. We found that a distal element of the promoter (-103 to -48) confers this cell-type-specific behavior through a mechanism in which the promoter activity is repressed in HeLa and SK-Hep1 cells but increased in Alexander cells. By using an inhibitor of protein synthesis, we obtained evidence that a labile repressor(s) confers the negative effect in SK-Hep1 cells. We also found an enhancerlike activity associated with a small DNA segment of the S promoter (-27 to + 30). This proximal element was active in HeLa and SK-Hep1 cells only in the absence of the distal negative element. Finally, analysis of S promoter deletion mutants demonstrated that the -27 to -17 region of the S promoter is crucial for its activity.
Mol Cell Biol 1988 Jun
PMID:The S promoter of hepatitis B virus is regulated by positive and negative elements. 316 91

We have investigated the role of liver-specific trans-acting factor(s) in the regulation of hepatitis B virus (HBV) gene expression. A recorder plasmid (pEcoAluCAT; HBV nucleotides 1 through 1878) was constructed containing the HBV enhancer and the promoter region of the pregenomic RNA, which was ligated to the bacterial chloramphenicol acetyltransferase (CAT) gene. Upon transfecting this plasmid into various cell lines, the CAT gene was expressed only in cells of liver origin. Moreover, competition cotransfections with pEcoAluCAT and plasmids containing HBV enhancer sequences in human hepatoblastoma-derived HepG2 cells indicated the presence of titratable trans-acting factor(s) in these cells. Gel mobility shift assays using HBV enhancer and core promoter domains confirmed the existence of sequence-specific DNA-binding proteins in liver cell nuclear extract which bound to these regions. These binding sites encompass 17- and 12-nucleotide palindromes in the HBV enhancer and core promoter domains, respectively, when mapped by the methylation interference assay.
Mol Cell Biol 1988 Dec
PMID:Identification of protein-binding sites in the hepatitis B virus enhancer and core promoter domains. 324 51

We describe the in vivo production of 5-bromodeoxyuridine- (5-BUdR) labelled M13 DNA by a thymine-requiring Escherichia coli strain. We show that the 5-BUdR-labelled M13 single-stranded DNA is not extruded into the culture medium, but accumulates inside the bacterial cells. On the basis of this observation, a procedure involving FPLC gel filtration already reported and used for the isolation of plasmid DNA has been adapted for the isolation of at least 90% pure 5-BUdR-labelled single-stranded DNA. An M13 probe, containing part of the Hepatitis B Virus (HBV) genome was constructed, and the corresponding 5-BUdR-labelled single-stranded DNA was used in hybridization experiments to detect homologous HBV target DNA. Picogram amounts (10(-19) moles) of the probe itself or the target DNA could be detected, by monoclonal anti-5-BUdR antibodies in an immunoenzymatic assay.
Mol Cell Probes 1987 Mar
PMID:5-Bromodeoxyuridine in vivo labelling of M13 DNA, and its use as a non-radioactive probe for hybridization experiments. 333 Nov 68

We have analyzed a series of plasmids in which the sequences located upstream from the hepatitis B virus (HBV) X gene were linked to the chloramphenicol acetyl transferase (CAT) gene. Expression of the marker CAT gene in transfected cells clearly demonstrated that sequences preceding the X gene contain an active promoter. RNA mapping by primer extension indicated that the RNA encoded by the X gene promoter initiates at multiple sites spanning nucleotides 1250 to 1350 on the HBV genome. Deletion within the adjacent HBV enhancer element region significantly reduced the activity of the X gene promoter, suggesting that the X gene promoter requires the enhancer element for maximal activity.
Mol Cell Biol 1987 Jan
PMID:Identification of a promoter element located upstream from the hepatitis B virus X gene. 349 93

Small spherical particles produced in the non-permissive phase of hepatitis B virus infection, when the viral genome is integrated into the chromosome of hosts, are rich in the product of the S gene, but poor in the product of the pre-S2 region. For the purpose of adding immunogenicity to spherical particles deficient in the pre-S2 region product, they were conjugated with a synthetic peptide of 19 amino acid residues. The peptide reproduced a hydrophilic area of the pre-S2 region product encoded by viral genomes of subtypes adr, ayw and ayr. The spherical particles supplemented with the pre-S2 peptide raised antibody to the pre-S2 region product in mice, in addition to antibody to the product of the S gene. Antibody to pre-S2 region product, prepared from sera of immunized mice by absorption with the S gene product, bound to spherical particles bearing pre-S2 region product, irrespective of adr, adw, ayw or ayr subtype, and agglutinated hepatitis B virions in immune electron microscopy. Based on the results obtained, the synthetic peptide may prove useful in adding protective efficacy to small spherical particles poor in pre-S2 region product.
Mol Immunol 1987 May
PMID:A synthetic peptide coded for by the pre-S2 region of hepatitis B virus for adding immunogenicity to small spherical particles made of the product of the S gene. 365 94

An immune response to epitopes localized on the preS region of the hepatitis B virus (HBV) envelope (env) is elicited during recovery from HBV infection and appears to play a role in virus clearance. Anti-preS antibodies (Ab) are expected to be protective against HBV infection as indicated by the virus-neutralizing capacity of Ab to a preS2-specific synthetic peptide preS(120-145). However, there is considerable amino acid variability between preS regions corresponding to distinct serological subtypes of HBV, raising the question whether the preS sequences are sufficiently related immunologically to have the potential of inducing cross-protective immunity. To answer this question concerning the preS2 region, antisera to synthetic peptides preS(120-153) and preS(128-153) corresponding to subtypes adw2 and ayw, as well as to the native env [ayw] protein were raised. Using the resulting polyclonal Ab, an immunological relatedness between preS2 sequences of subtypes adw2 and ayw was demonstrated. On the other hand, Ab selected by affinity chromatography or by cloning hybridoma cells may recognize with strong preference subtype-specific determinants within the preS2 region when the compared antigens are in solution rather than on the solid phase. These findings have implication for the design of: (1) preS2-specific immunogens and (2) immunoassays for quantitation of preS2 sequences in HBV env proteins.
Mol Immunol 1987 Jun
PMID:Immunological cross-reactivity between preS2 sequences of the hepatitis B virus envelope proteins corresponding to serological subtypes adw2 and ayw. 365 96


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