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)

Equine herpesvirus type 1 (EHV-1) strain Ab4 gene 67 has no counterpart in any herpesvirus sequenced to date. To identify and characterize the product of EHV-1 gene 67, we have expressed the putative amino acids 11 to 260 encoded by gene 67 as a beta-galactosidase fusion protein in Escherichia coli. The expressed fusion protein has been used to generate an antiserum raised against the gene 67 product. Immunoblotting and immunoprecipitation experiments have revealed that the anti-67 serum specifically recognizes a polypeptide with an M(r) of 36,000 (the 36K polypeptide) in infected cell extracts. The gene 67 protein is regulated as an early polypeptide in EHV-1 strain Ab4 infected cells and post-translational modification experiments have revealed that the protein is phosphorylated, but not glycosylated. The gene 67 protein has been transiently expressed in BHK-21/C13 cells using plasmid pCMV67, which contains the putative gene 67 ORF under the control of the cytomegalovirus immediate early promoter. Immunoblotting experiments with anti-67 have shown that the 36K protein is expressed at high levels in transfected cells. From both immunofluorescence and cellular fractionation experiments it is concluded that the gene 67 protein is associated with intracellular membranes and produces novel ribbon or filament-like structures within the cytoplasm of infected cells. We have demonstrated that the gene 67 product is a component of the virion nucleocapsid/tegument.
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PMID:Identification and characterization of the protein product of gene 67 in equine herpesvirus type 1 strain Ab4. 789 46

AGA and AGG codons for arginine are the least used codons in Escherichia coli. Previous findings have shown that these codons are used preferentially within the first 25 codons in E. coli genes. More than 100 genes having a single AGA/AGG codon within the first 25 codons were identified to be associated with various essential cellular functions. The lacZ gene, containing 5 AGG codons after the tenth codon from the initiation codon, was constructed as a model system. The production of beta-galactosidase was inhibited almost completely during the stationary phase, whereas the production of the control beta-galactosidase without AGG codons was not. The inhibitory effect by the 5 AGG codons was substantially suppressed either by coexpressing the argU gene for tRNA(ArgUCU/CCU) or by moving the 5 AGG codons by > 50 codons away from the initiation codon. In addition, the production of a number of proteins resolved by two-dimensional gel electrophoresis was enhanced significantly during the stationary phase in the cells harboring a plasmid containing argU. At least one of them was identified as the hns product encoded by an ORF having an AGA codon at the nineteenth position. On the basis of these results, it is proposed that the expression of a group of essential genes for various cellular functions that have a single AGA/AGG codon very close to the initiation codon are globally regulated by the availability of the least abundant tRNA(ArgUCU/CCU). A model for this regulation is proposed.
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PMID:Role of the AGA/AGG codons, the rarest codons in global gene expression in Escherichia coli. 795 22

Feline herpesvirus type 1 (FHV-1) mutants were constructed, carrying a beta-galactosidase marker gene integrated into the region downstream of the gene encoding the homologue of glycoprotein C (gC) of herpes simplex virus type 1. In cell culture, no differences in replication were observed between mutants and the parent FHV-1 strain. However, in experimentally infected cats, mutants caused fewer clinical signs after oronasal administration although they replicated to the same extent as the parental strain. Sequence analysis in the region of the UL segment surrounding the insertion site revealed an open reading frame (ORF 2) encoding a putative polypeptide of 21K. RNA analysis indicated a corresponding transcript of 0.8 kb that was detected late after infection of cells in culture. This particular UL locus downstream of the gC gene has not been thoroughly investigated in any of the herpesviruses. The putative gene product showed only limited evolutionary conservation since similarity could be found only with the assumed homologue of equine herpesvirus type 1. Further characterization of this newly identified FHV-1 gene involved in virulence may provide insight into the development of disease owing to herpesvirus infection.
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PMID:The gene downstream of the gC homologue in feline herpes virus type 1 is involved in the expression of virulence. 796 20

We have marked a Drosophila transposable element--the LINE-like I element--with an intron-containing indicator gene inserted in place of a large deletion in the I element second ORF encompassing the reverse transcriptase domain, and this marked element was placed downstream to a potent actin promoter. An expression vector for the I element ORFs was also constructed, under the same heterologous promoter. The indicator gene contains a lacZ reporter gene the expression of which is conditioned by retrotransposition of the marked element, thus allowing detection of transposition events by testing for either beta-galactosidase expression or occurrence of spliced DNA molecules. The marked I element was introduced into Drosophila melanogaster cells in culture by transfection. Spliced DNA copies of the marked element and specifically stained beta-galactosidase-expressing cells were detected only upon co-transfection with the I expression vector, thus indicating that an ORF2-deleted element can be complemented in trans for transposition. This simple assay for retrotransposition in Drosophila cells in culture provides a tool for the rapid analysis of the mechanism of I transposition in its cis and trans sequence requirements.
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PMID:Retrotransposition of a marked Drosophila line-like I element in cells in culture. 819 Jun 41

Polyamine stimulation of the synthesis of oligopeptide-binding protein (OppA) was shown to occur mainly at the level of translation by measuring OppA synthesis and its mRNA level. Several artificial oppA genes were constructed by site-directed mutagenesis. These synthesize different kinds of OppA mRNAs: mRNAs differing in the size of 5'-untranslated region; mRNAs having the Shine-Dalgarno (SD) sequence in a different position; mRNAs having different secondary structure in the region of the SD sequence; and fusion mRNAs consisting of the 5'-untranslated region of OppA mRNA and the open reading frame of beta-galactosidase. By measuring the synthesis of OppA or beta-galactosidase from these mRNAs, we found that the 171-nucleotide 5'-untranslated region and 145 nucleotides of the ORF of OppA mRNA are involved in the polyamine stimulation of OppA synthesis. When the secondary structure of the above region of OppA mRNA was analyzed by optimal computer folding, it was shown that the degree of polyamine stimulation of OppA protein synthesis was dependent on the structure of the SD sequence in addition to its position. Loose base pairing of the SD sequence with other regions of the mRNA caused strong polyamine stimulation, while intense base pairing of the SD sequence with other regions of the mRNA resulted in insignificant or weak polyamine stimulation.
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PMID:Molecular mechanism of polyamine stimulation of the synthesis of oligopeptide-binding protein. 902 Jan 14

We summarize in this communication the data supporting the two functions of ribosome recycling factor (RRF, originally called ribosome releasing factor). The first described role involves the disassembly of the termination complex which consists of mRNA, tRNA and the ribosome bound to the mRNA at the termination codon. This process is catalyzed by two factors, elongation factor G (EF-G) and RRF. RRF stimulated protein synthesis as much as eight-fold in the in vitro lysozyme synthesis system, when ribosomes were limiting. In the absence of RRF, ribosomes remain mRNA-bound at the termination codon and translate downstream codons. In the in vitro system, the site of reinitiation is the triplet codon 3' to the termination codon. RRF is an essential protein for bacterial life. Temperature sensitive (ts) RRF mutants were isolated and in vivo translational reinitiation due to inactivation of ts RRF was demonstrated using the beta-galactosidase reporter gene placed downstream from the termination codon. A second function of RRF involves preventing errors in translation. In polyphenylalanine synthesis programmed by polyuridylic acid, misincorporation of isoleucine, leucine or a mixture of amino acids was stimulated upto 17-fold when RRF was omitted from the in vitro system. RRF did not influence the large error (10-fold increase) induced by streptomycin. This means that RRF participates not only in the disassembly of the termination complex but also in peptide elongation. Extending this concept and its conventional role for releasing ribosomes from mRNA, involvement of RRF in the reinitiation in the 3A' system (a construct using S aureus protein A, a collaborative work with Dr Isaksson), in programmed frame shifting, in trans-translation with 10Sa RNA (collaborative work with Dr Muto), and in the reinitiation downstream from the ORF A of the IS 3 (insertion sequence of a transposon, collaborative work with Dr Sekine) are discussed on the basis of preliminary data to be published elsewhere. Finally, we review the known RRF sequences from various organisms including eukaryotes and discuss the possible mechanism for disassembly of the eukaryotic termination complex.
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PMID:Dual functions of ribosome recycling factor in protein biosynthesis: disassembling the termination complex and preventing translational errors. 915 Aug 73

ORF slr0798, now designated ziaA, from Synechocystis PCC 6803 encodes a polypeptide with sequence features of heavy metal transporting P-type ATPases. Increased Zn2+ tolerance and reduced 65Zn accumulation was observed in Synechococcus PCC 7942, strain R2-PIM8(smt), containing ziaA and upstream regulatory sequences, compared with control cells. Conversely, reduced Zn2+ tolerance was observed following disruption of ziaA in Synechocystis PCC 6803, and ziaA-mediated restoration of Zn2+ tolerance has subsequently been used as a selectable marker for transformation. Nucleotide sequences upstream of ziaA, fused to a promoterless lacZ gene, conferred Zn2+-dependent beta-galactosidase activity when introduced into R2-PIM8(smt). The product of ORF sll0792, designated ZiaR, is a Zn2+-responsive repressor of ziaA transcription. Reporter gene constructs lacking ziaR conferred elevated Zn2+-independent expression from the ziaA operator-promoter in R2-PIM8(smt). Gel retardation assays detected ZiaR-dependent complexes forming with the zia operator-promoter and ZiaR-DNA binding was enhanced by treatment with a metal-chelator in vitro. Two mutants of ZiaR (C71S/C73S and H116R) bound to, and repressed expression from, the ziaA operator-promoter but were unable to sense Zn2+. Metal coordination to His-imidazole and Cys-thiolate ligands at these residues of ZiaR is thus implicated in Zn2+-perception by Synechocystis PCC 6803.
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PMID:An SmtB-like repressor from Synechocystis PCC 6803 regulates a zinc exporter. 972 72

A 5 kb region upstream of katA at 82 degrees on the Bacillus subtilis chromosome contains five ORFs organized in an operon-like structure. Based on sequence similarity, three of the ORFs are likely to encode an ABC transport system (ssuBAC) and another to encode a monooxygenase (ssuD). The deduced amino acid sequence of the last ORF (ygaN) shows no similarity to any known protein. B. subtilis can utilize a range of aliphatic sulfonates such as alkanesulfonates, taurine, isethionate and sulfoacetate as a source of sulfur, but not when ssuA and ssuC are disrupted by insertion of a neomycin-resistance gene. Utilization of aliphatic sulfonates was not affected in a strain lacking 3'-phosphoadenosine 5'-phosphosulfate (PAPS) sulfotransferase, indicating that sulfate is not an intermediate in the assimilation of sulfonate-sulfur. Sulfate or cysteine prevented expression of beta-galactosidase from a transcriptional ssuD::lacZ fusion. It is proposed that ssuBACD encode a system for ATP-dependent transport of alkanesulfonates and an oxygenase required for their desulfonation.
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PMID:Bacillus subtilis genes for the utilization of sulfur from aliphatic sulfonates. 978 4

We have cloned and characterized the MCM16 gene required for the maintenance of minichromosomes in the yeast Saccharomyces cerevisiae. This gene corresponds to a 181-amino acid ORF, YPR046W, on chromosome XVI. Mutant cells carrying minichromosomes accumulate them in higher copy numbers than do wild-type cells. Intact dicentric plasmid could be recovered from the mutant, in contrast to the wild-type, in which the plasmid suffered frequent deletions. A wild-type centromere, CEN6, acts as a block to the transcription of a reporter gene, such as beta-galactosidase. This block was less effective in the mutant than in the wild-type strain, suggesting alterations in kinetochore assembly in the former. The mutant also showed increased sensitivity to the antimitotic drugs benomyl and thiabendazole. The mcm16 mutation caused a high rate of loss of chromosome III, without any significant increase in the recombination frequency. A strain carrying a deletion-disruption derivative of the MCM16 gene was viable and, when compared to the wild-type, did not show any significant changes in growth rate or cell morphology at 16, 23 and 37 degrees C. These properties show that MCM16 is required for an important but nonessential role that governs the kinetochore-microtubule mediated process of chromosome segregation.
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PMID:The MCM16 gene of the yeast Saccharomyces cerevisiae is required for chromosome segregation. 986 78

Attempts were made to linearize the DNA of Choristoneura fumiferana (Cf) multicapsid nucleopolyhedrovirus (MNPV), in order to improve the efficiency of generation of recombinant viruses after transfection. A unique site for the restriction enzyme Sse83871 was found in ORF p48. The requirement for this ORF during virus replication was investigated by molecular analyses including sequencing, transcriptional analysis and inactivation by insertion of marker genes. Sequence analysis showed that ORF p48 consists of 1233 nucleotides encoding a potential protein of 47.88 kDa. The proteins encoded by ORF p48 from CfMNPV and Orgyia pseudotsugata MNPV contain 411 amino acids while that from CfDEFNPV (a virus that is defective for infection by the per os route) is slightly smaller, at 408 amino acids. Transcriptional and primer extension analyses showed that the mRNA is initiated from a typical baculovirus late gene ATAAG motif. The mRNA was detected at 24 h post-infection (p.i.), reached maximum levels at 48 h p.i. and declined by 96 h p.i., which confirmed the late property of the gene. Inactivation of the gene was attempted by inserting a cassette containing either the gene encoding beta-galactosidase or that encoding green fluorescent protein. Blue or fluorescent green plaques of infected cells were observed after transfection. Attempts to generate a plaque-purified virus were not successful. Restriction enzyme analysis showed that the marker genes were inserted randomly at positions other than the p48 locus. This indicated that the gene may be needed for virus replication. The gene is relatively well conserved among baculoviruses but its function remains unclear.
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PMID:Molecular analysis of the p48 gene of Choristoneura fumiferana multicapsid nucleopolyhedroviruses CfMNPV and CfDEFNPV. 1042 53


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