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)

In Saccharomyces cerevisiae cells harboring a GAL1 promoter-linked beta-galactosidase gene, the simultaneous expression of Escherichia coli DNA topoisomerase I and inactivation of yeast DNA topoisomerases I and II reduces the cellular level of beta-galactosidase to an undetectable level. Analysis of intracellular mRNA level and the density of RNA polymerase along DNA indicates that this reduction is due to the suppression of transcription and that both plasmid-borne and chromosomally located genes are affected. These results are interpreted in terms of inhibition of transcription in vivo due to positive supercoiling of the DNA template: preferential removal of transcription-generated negative supercoils by E. coli DNA topoisomerase I in the absence of both yeast DNA topoisomerases I and II results in the accumulation of positive supercoils in intracellular DNA. In normal prokaryotic or eukaryotic cells, accumulation of positive supercoils is presumably avoided through the balanced actions of DNA topoisomerases.
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PMID:Positive supercoiling of DNA greatly diminishes mRNA synthesis in yeast. 133 10

Mutations in top, the structural gene for Escherichia coli DNA topoisomerase I, have been identified and mapped at 28 min on the chromosome, near cysB. Strains carrying deletions of the top gene are viable. The top mutations, however, do exert pleiotropic effects on transcription and transposition. Mutants lacking DNA topoisomerase I have a more rapid rate of induction and a higher level of catabolite-sensitive enzymes including tryptophanase and beta-galactosidase. This general activation of transcription by top mutations can be attributed to an increase in the negative superhelicity of the DNA in vivo when the topoisomerase activity is abolished. The frequency of transposition of Tn5, a transposon carrying kanamycin resistance, is decreased by a factor of 40 or more in top mutants. A direct or indirect role of the topoisomerase in transposition is discussed. The transposition frequency of Tn3, however, is not dependent on top. Based on the studies of the E. coli top mutants, it appears that the supX gene, which was originally studied in Salmonella typhimurium [Dubnau, E. & Margolin, P. (1972) Mol. Gen. Genet. 117, 91-112] is likely to be the structural gene for DNA topoisomerase I.
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PMID:Mutations in the gene coding for Escherichia coli DNA topoisomerase I affect transcription and transposition. 626 7

DNA topoisomerase I (topo I) from Drosophila melanogaster contains a nonconserved, hydrophilic N-terminal domain of about 430 residues upstream of the conserved core domains. Deletion of this N terminus did not affect the catalytic activity of topo I, while further removal of sequences into the conserved regions inactivated its enzymatic activity. We have investigated the cellular function of the Drosophila topo I N-terminal domain with top1-lacZ transgenes. There was at least one putative nuclear localization signal within the first 315 residues of the N-terminal domain that allows efficient import of the large chimeric proteins into Drosophila nuclei. The top1-lacZ fusion proteins colocalized with RNA polymerase II (pol II) at developmental puffs on the polytene chromosomes. Either topo I or the top1-lacZ fusion protein was colocalized with RNA pol II in some but not all of the nonpuff, interband loci. However, the fusion proteins as well as RNA pol II were recruited to heat shock puffs during heat treatment, and they returned to the developmental puffs after recovery from heat shock. By immunoprecipitation, we showed that two of the largest subunits of RNA pol II coprecipitated with the N-terminal 315-residue fusion protein by using antibodies against beta-galactosidase. These data suggest that the topo I fusion protein can be localized to the transcriptional complex on chromatin and that the N-terminal 315 residues were sufficient to respond to cellular processes, especially during the reprogramming of gene expression.
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PMID:Targeting to transcriptionally active loci by the hydrophilic N-terminal domain of Drosophila DNA topoisomerase I. 963 19

Linear expression constructs can facilitate gene function studies. We describe a method to generate linear expression constructs for mammalian cells by one-step polymerase chain reaction (PCR) with vaccinia DNA topoisomerase I (TOPO). Cytomegalovirus (CMV) 5\' promoter, the gene of interest, and V5 bovine growth hormone (BGH) polyA 3\' terminator elements were PCR-amplified with target-specific primers containing vaccinia DNA TOPO-specific sequence and complementary sequence to each other. We amplified specific and complementary sequences. These three elements were directionally joined with vaccinia TOPO. The joined products were then directly transfected into Chinese hamster ovary cells. Compared with the transfection of supercoiled plasmids, comparable expression signals were obtained for green fluorescent protein, chloramphenicol acetyltransferase, and beta-galactosidase proteins using Western blots. This is a quick and efficient method to generate linear expression constructs. Unlike Invitrogen TOPO Tools, our method avoided the secondary round of PCR and more rapidly yielded correct joining products. This method can be easily used in the function test of uncharacterized open reading frames.
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PMID:Generation of linear expression constructs by one-step PCR with vaccinia DNA topoisomerase I. 1740 Nov 45