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)

The binding sites for human interferon-alpha (IFN-alpha) have been characterized on human lymphoblastoid, melanoma, rhabdomyosarcoma, and cervical carcinoma cells. Crosslinking of iodinated-recombinant DNA-derived IFN-alpha-Con1, an analog of the known IFN-alpha subtypes, to the cell surface with disuccinimidyl suberate yielded four IFN-receptor complexes of 118, 138, 159, and 260 kD on all cell lines that specifically bind IFN-alpha. Since IFN-alpha exists in solution as monomers, dimers, and trimers, and the three lower molecular weight IFN-alpha-receptor complexes differ by the molecular weight of IFN-alpha (20 kD), this suggests that the human IFN-alpha receptor of 100 kD binds more than one molecule of IFN-alpha. The higher molecular weight complex of 260 kD may result from dimerization of the receptor. None of these complexes was observed in a rhabdomyosarcoma subclone that does not specifically bind IFN-alpha. Pretreatment of cells with trypsin abolished the formation of these complexes. Pretreatment of cells with neuraminidase did not reduce IFN-alpha binding, but increased the electrophoretic mobility of all four IFN-alpha-receptor complexes. Other glycosidases (i.e., mannosidase, beta-galactosidase, and endoglycosidase F) had no effects on IFN-alpha binding or mobility of complexes. Thus, although the IFN-alpha receptor is a glycoprotein, the glycosylated portion is apparently not part of the IFN-alpha-binding domain. The formation of IFN-alpha-receptor complexes is independent of the duration of incubation with IFN (from 5 min to 1 h at 15 degrees C).(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Characterization of interferon-alpha binding sites on human cell lines. 246 92

The fusion of the N-terminal 461 bp of the human interferon-alpha 2 (INF) in frame to the beta-galactosidase gene from Escherichia coli is described. The presence of the expected DNA sequence was shown by restriction mapping and DNA sequencing. A fusion protein was demonstrated in crude extracts of E. coli by Western blots using polyclonal anti-beta-galactosidase and monoclonal anti-IFN antibodies. Using monoclonal antibodies specific for the N-terminal region of IFN-alpha and cell-free extracts from an E. coli strain containing the fusion protein, we set up a simple competitive enzyme-linked immunosorbent assay for human interferon. The test described here was linear down to a lower detection limit of at least 1000 Units, or 5 ng human IFN.
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PMID:A simple competitive enzyme-linked immunosorbent assay using antigen-beta-galactosidase fusions. 311 82

The complete nucleotide sequence of a human interferon-alpha gene is reported. The gene, designated IFN-alpha M1, was isolated from a human genomic library in phage lambda Charon 4A using synthetic oligonucleotides as hybridization probes. Based on a comparison of nucleotide sequence data obtained from this recombinant phage with published interferon-alpha gene sequences, a region of DNA capable of coding for a pre-interferon of 189 amino acids was identified. An AluI fragment containing the coding region for the mature interferon was inserted into the HincII site of the phage M13mp11, resulting in a fusion of portions of the IFN-alpha M1 and the beta-galactosidase genes. Antiviral activity was detected in extracts from E. coli infected with the recombinant M13 phage carrying the fused gene. The antiviral activity was completely neutralized by antibodies to human interferon-alpha.
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PMID:Nucleotide sequence and expression in E. coli of a human interferon-alpha gene selected from a genomic library using synthetic oligonucleotides. 608 30

The development and utilization of a tissue culture system for the analysis of quiescent, nonreplicating herpes simplex virus type 1 (HSV-1) genomes is described. It was demonstrated previously that the HSV-1 Vmw65 mutant in1814, which is impaired for immediate early (IE) transcription, was retained for many days in human fetal lung (HFL) fibroblasts in a quiescent 'latent' state. Molecular analysis of the viral genome was not possible, however, due to residual expression of IE proteins and consequent cytotoxicity at high m.o.i. In the study reported here, IE transcription was reduced further by pretreatment of cells with interferon-alpha (IFN-alpha) and by the use of mutant in1820, a derivative of in1814 in which the Vmw110 promoter was replaced by the Moloney murine leukaemia virus (Momulv) enhancer. The Momulv enhancer was not expressed under IE conditions; thus in1820 was more impaired for replication than in1814 and behaved as if deficient for both Vmw65 and Vmw110. In cells pretreated with IFN-alpha and subsequently infected with in1820 cytotoxicity was overcome, enabling a tissue culture system to be developed in which all cells stably retained at least one quiescent viral genome. To assist the analysis of gene expression, in1820 was further modified by insertion of the Escherichia coli lacZ gene controlled by the human cytomegalovirus enhancer (mutant in1883) or the HSV-1 immediate early Vmw110 promoter (in1884). Expression of beta-galactosidase was not detected after infection of IFN-alpha-pretreated cells with in1883 or in1884 but could be induced in almost all cells containing a viral genome, by superinfection of cultures. In1820-derived viruses were retained for at least 9 days and were not reactivated by subculture of cells. A regular arrangement of nucleosomes, as found in cellular chromatin, was not detected on the viral genome at the thymidine kinase locus. The non-linear genome was a template for reactivation with no requirement for prior conversion to a linear form. A small number of remaining linear genomes resulted from incomplete uncoating of input virus.
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PMID:Quiescent viral genomes in human fibroblasts after infection with herpes simplex virus type 1 Vmw65 mutants. 778 70

A genetic system to display proteins as their active and functional forms on the cell surface of yeast, Saccharomyces cerevisiae, has been exploited. Surface-engineered (arming) cells displaying amylase or cellulase could assimilate starch or cellulose as the sole carbon source, although S. cerevisiae can not intrinsically assimilate them. Arming cells with a green fluorescent protein (GFP) from Aequorea victoria can emit green fluorescence from the cell surface in response to the environmental conditions. From these results, we attempted to construct a system to monitor the foreign protein production in yeast by simultaneous displaying the enhanced GFP (EGFP). The expression in yeast of the Escherichia coli beta-galactosidase-encoding gene was examined as an example of intracellular production and that of the human interferon-alpha (omega, IFN-omega)-encoding gene as an example of extracellular production. Their productions and the simultaneous surface-display of EGFP as a reporter were controlled by the same promoter, GAL1. The relationship among fluorescence signals and their productions was evaluated. The surface-display system, unlike one using tag-proteins, would be able to facilitate the monitoring of native protein productions in bioprocesses using living cells in real time by the combination of promoters and GFP variants.
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PMID:Development of combinatorial bioengineering using yeast cell surface display--order-made design of cell and protein for bio-monitoring. 1456 12