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 coding region of the chicken histone H1.03 gene was cloned to a bacterial expression vector, and the 291-amino acid H1-beta-galactosidase fusion protein was isolated after induction with IPTG. The fusion protein recognizes the 5'-TTGGCAnnnTGCCAA-3' motif on DNA. The H1 globular domain was initially shown to be responsible for the sequence-specific binding by functional deletion analysis. This function may be indispensable for the role of H1 as a determinant of nucleosome positioning and as a eukaryotic repressor.
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PMID:The histone H1-lacZ' fusion protein produced in Escherichia coli binds to the 5'-TTGGCAnnnTGCCAA-3' motif on DNA. 189 49

The yeast Cdc7 protein is indispensable to initiation of nuclear DNA replication, based on the phenotype of the conditional, temperature-sensitive (ts) cdc7 mutants at the restrictive temperature. This protein has likewise been implicated in commitment to meiotic DNA recombination and induced mutagenesis, which may result from error-prone DNA repair. Our previous work revealed sequence similarity between the Cdc7 protein and known protein kinases. To determine whether it possesses kinase activity, we have immunoprecipitated the protein from Cdc7-overproducing yeast cells by using polyclonal antibodies raised against a nondenatured beta-galactosidase-Cdc7 fusion protein. In this report, we demonstrate that Cdc7 immune complexes are capable of phosphorylating mammalian histone H1 on serine and/or threonine residues. Immune complexes derived from cells harboring the cdc7-2 ts mutant gene on a high copy number plasmid possess a thermolabile kinase activity. Thus, we postulate that Cdc7 may regulate the various DNA metabolic pathways by phosphorylating one or more target substrates. Because Cdc7 kinase acts downstream of Cdc28/cdc2 kinase function at "start," the transition from G1 to S phase in the cell cycle may be the result of a cascade of protein phosphorylation.
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PMID:DNA metabolism gene CDC7 from yeast encodes a serine (threonine) protein kinase. 216 54

The interrelationship between autoepitopes, DNA-binding domains, and C-reactive protein (CRP)-binding domains on a histone H1 molecule was examined using fusion proteins of beta-galactosidase and truncated histone H1 molecules. At least two CRP-binding sites were detected on a histone H1 molecule. Site 1 was composed of approximately 25 amino acids and calcium ion was required for the binding of CRP. Site 2, composed of approximately 20 amino acids and not requiring calcium ion, was identical or located very close to a DNA-binding domain and an epitope of anti-histone H1 autoantibodies in SLE sera. These data suggest that, at physiological ionic strength, histone H1 of either free or immune-complexed form could bind to CRP via site 1. These data are discussed with respect to the possible role of CRP in the handling and clearance of immune complexes in patients with systemic lupus erythematosus.
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PMID:Interrelationship between autoepitope, DNA-binding domain, and CRP-binding domain on a histone H1 molecule. 767 45

We have examined the central nervous system (CNS) of developing and adult transgenic mice carrying sequences upstream of the histone H1 zero gene fused to the E. coli beta-galactosidase gene (lac Z). The transgene is induced in a subset of the neuronal population during postnatal development, coinciding with neuronal terminal differentiation. At postnatal day 9, the earliest time at which the transgene product can be detected, positive neurons are observed in the granular layer of the cerebellar cortex and in the pyramidal fields of the hippocampus. The transgene is then induced in other areas of the CNS, such as the neocortex, thalamus, hypothalamus, olfactory bulb, globus pallidus superior and inferior colliculus, substantia nigra, pontine nuclei and brain stem. Induction is unrelated with determination and quiescence, which are essentially prenatal. The overlapping of the temporal and regional patterns of transgene activity with those of the endogenous protein shows that the accumulation of H1 zero in differentiating neurons is at least in part under transcriptional control. In the light of these results, the H1 zero gene appears as the only mammalian histone gene that specifically responds to terminal differentiation. However, not all terminally differentiated neurons express H1 zero at detectable levels. For instance, Purkinje cells are negative. In neurons, terminal differentiation appears thus as a necessary, but not a sufficient condition for increased H1 zero expression.
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PMID:Transcriptional activation of histone H1 zero during neuronal terminal differentiation. 795 58

A recombinant histone (NLS-H1) containing both the SV40 large T antigen nuclear localization signal and the carboxy-terminal domain of human histone H1(0) was produced in bacteria. NLS-H1-plasmid DNA complexes, in the presence of chloroquine, mediated reporter gene transfer into cultured cells with similar efficiencies as plasmid DNA-cationic lipid (lipofectin) complexes. NIH-3T3 or COS-7 cells transfected with NLS-H1-plasmid DNA-lipofectin complexes expressed at least 20 times more luciferase or had at least 2.5 times more beta-galactosidase-positive cells than those transfected with plasmid DNA-lipofectin complexes. Foreign gene expression was also improved by other DNA-binding proteins and cationic lipid formulations, yet the greatest enhancement was obtained with complexes containing either NLS-H1 or calf thymus histone H1. Histone H1-plasmid DNA-lipofectin complexes were internalized by a greater number of cells than plasmid DNA-lipofectin complexes.
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PMID:Gene transfer into mammalian cells using histone-condensed plasmid DNA. 884 98

Ligand-mediated targeting of DNA was validated by condensing a plasmid DNA encoding the beta-galactosidase (beta-gal) gene with a basic fibroblast growth factor (FGF2) that was first chemically conjugated to polylysine (K). The conditions that gave optimal binding of this FGF2 to DNA also generated the highest level of beta-gal expression when added to FGF2 target cells like COS-1, 3T3, baby hamster kidney (BHK), or endothelial cells. This beta-gal activity increased in a time- and dose-dependent manner and was dependent on the inclusion of FGF2 in the complex. FGF receptor specificity was demonstrated by competition of the complex with FGF2 and heparin, and by the failure of cytochrome c or histone H1 to mimic the gene-targeting effects of FGF2. The expression of beta-gal was also endosome dependent because chloroquine increased beta-gal expression 8-fold and endosome disruptive peptides increased expression of beta-gal 26-fold. Taken together these findings establish that DNA can be introduced into cells through the high affinity FGF receptor complex, and while its efficiency will require significant enhancements to achieve sustained and elevated transgene expression, the possibility that the technique could be used to deliver DNAs encoding cytotoxic molecules is discussed.
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PMID:Targeting DNA to cells with basic fibroblast growth factor (FGF2). 896 34

The existence of histone H1 in the yeast, Saccharomyces cerevisiae, has long been debated. In this report we describe the presence of histone H1 in yeast. YPL127c, a gene encoding a protein with a high degree of similarity to histone H1 from other species was sequenced as part of the contribution of the Montreal Yeast Genome Sequencing Group to chromosome XVI. To reflect this similarity, the gene designation has been changed HHO1 (Histone H One). The HHO1 gene is highly expressed as poly A+ RNA in yeast. Although deletion of this gene had no detectable effect on cell growth, viability or mating, it significantly altered the expression of beta-galactosidase from a CYC1-lacZ reporter. Fluorescence observed in cells expressing a histone H1-GFP protein fusion indicated that histone H1 is localized to the nucleus.
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PMID:Histone H1 in Saccharomyces cerevisiae. 904 96

Ternary complexes of plasmid DNA, histone H1 protein and amphipathic polyamines (PAPA) were able to mediate the efficient transfection of 3T3. HeLa and COS cells in culture. Using both the beta-galactosidase and luciferase reporter gene systems, the transfection efficiency of PAPA complexes was comparable to that of DOSPA/PE cationic liposomes, considered to be a highly-efficient transfection reagent. Using three different assays of cellular toxicity (propidium iodide, BCECF-AM and Trypan Blue), the PAPA complexes caused minimal cellular toxicity. These results indicate that PAPA complexes are useful transfection reagents for the study of gene expression and function in cultured cells.
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PMID:Protein/amphipathic polyamine complexes enable highly efficient transfection with minimal toxicity. 923 46

We have investigated the nuclear transport of the replacement histone H1(0) and have searched for its nuclear localization sequence (NLS). The lysine-rich H1(0) histone differs from the other H1 histones with respect to its mode of expression and to the processing of the respective mRNA. Using the digitonin-permeabilized cell import assay we demonstrate that H1(0) is transported into the nucleus in an energy- and temperature-dependent manner. In competition experiments we show that the transport of H1(0) from the cytoplasm into the nucleus is competed by the SV40 T-antigen-NLS-peptide coupled to HSA, an established substrate of the importin pathway. In transfection studies we have expressed in HeLa cells a series of plasmid constructs containing different fragments of the coding region of the H1(0) histone gene that were fused to the beta-galactosidase gene, and we have determined the subcellular localization of each fusion protein. The results show that H1(0) contains multiple transport-competent sequence elements that can function as NLS and that H1(0) meets the requirements for a transport into the nucleus by an importin-dependent pathway.
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PMID:The histone H1(0) contains multiple sequence elements for nuclear targeting. 977 Mar 63

Tissue-specific gene transfer remains one of the main challenges to deliver genes into designated and/or disseminated cells. We have previously shown successful gene transfer with a nonviral gene delivery system based on the simple chemical conjugation of plasmid DNA with antibody. However, this approach was hampered by low efficiency due to the poor translocation rate of DNA to the nucleus. To improve this approach, we have modified our vector by introducing noncovalent binding between the antibody and DNA, allowing the possibility to introduce different important molecules. The noncovalent association was achieved with neutravidin and biotinylated components: (1) biotinylated antibodies; (2) a biotinylated hemagglutinin fusogenic peptide of influenza virus to favor endosomal escape; and (3) biotinylated histone H1 to compact, protect, and associate DNA to the complex. We report here that this delivery system can be internalized by tumor cells targeted by a specific monoclonal antibody, permits the protection of the transfected DNA, and allows its subsequent transfer into the nucleus after escape from the endosomal compartment. We also demonstrate that, in vitro, gene transfer with this vector showed much higher reporter activity in cells (15 vs. 0.5%) and a stronger production of murine interleukin 2 as compared with our previous vector. In vivo, a single intravenous injection of the vector containing an antibody directed to the G250 renal cell carcinoma-associated antigen led to beta-galactosidase expression in engrafted tumor bearing G250 but not in G250-negative tumor or in other tissues. Altogether, these results indicate that our antibody-based vector is suitable to promote gene delivery in vitro and in vivo in tumor cells.
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PMID:In vivo-targeted gene delivery using antibody-based nonviral vector. 1206 43


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