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:2.3.1.28 (chloramphenicol acetyltransferase)
5,100 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Transcobalamin II (TCII) is a plasma protein that binds vitamin B12 (cobalamin; Cbl) and facilitates the cellular uptake of the vitamin by receptor-mediated endocytosis. In genetic disorders that are characterized by congenital deficiency of TCII, intracellular Cbl deficiency occurs, resulting in an early onset of megaloblastic anemia that is sometimes accompanied by a neurologic disorder. To define the genetic basis for TCII deficiency, we have cloned and characterized the human gene that encodes this protein. The gene spans a minimum of 18 kbp and contains nine exons and eight introns, with a polyadenylation signal sequence located 509 bp downstream from the termination codon and a transcription initiation site beginning 158 bp upstream from the ATG translation start site. The 5' flanking DNA does not have a TATA or CCAAT regulatory element, but a 34-nucleotide stretch beginning just upstream of the CAP site contains four tandemly organized 5'-CCCC-3' tetramers. This sequence is a motif for a trans-active transcription factor (ETF) that regulates expression of the epidermal growth factor receptor gene (EGFR), which also lacks TATA and CCAAT regulatory elements. A GC-rich sequence that binds the SP1 protein is located 356 nucleotides upstream from the first of the series of CCCC tetramers. Although this GC sequence is at an unusual location with respect to the CAP site, a 507-bp fragment containing this GC box drives the chloramphenicol acetyltransferase (CAT) reporter gene after transient transfection into NIH 3T3 cells. No CAT activity was observed when a 420-bp fragment lacking this GC box but containing the ETF-binding domains was similarly transfected into this cell line. One consensus and two atypical motifs for the c-myc ligand are located downstream and upstream, respectively, of the GC box, and this could explain the elevated plasma TCII observed in some patients with multiple myeloma, as the c-myc product is overexpressed in some myeloma cells. Restriction endonuclease digestion of genomic DNA from eight normal subjects with Taq I, Hinfl, Msp I, and Bgl I identified three patterns of restriction fragment length polymorphism (RFLP). A number of the exon/intron splice junctions of human TCII, TCI, and IF genes are located in homologous regions of these proteins, providing evidence that these genes have evolved by duplication of an ancestral gene. This characterization of the TCII gene and the RFLP should facilitate the identification of the mutation(s) responsible for the genetic abnormalities of TCII expression.
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PMID:The cloning and characterization of the human transcobalamin II gene. 774 31

A total of 26 staphylococcal strains isolated from mink with urinary tract infections as well as from the environment of the mink were examined for antibiotic resistance and prevalence of plasmids mediating resistance to the antibiotics applied for prophylactic or therapeutic purposes. Chloramphenicol resistance (Cmr) which occurred in fourteen of the eighteen Staphylococcus lentus strains, but in none of the Staphylococcus intermedius and Staphylococcus xylosus strains, was shown to be mediated by small plasmids of 3.6 to 4.6 kb. On the basis of restriction endonuclease mapping and hybridization experiments, four different types of Cmr plasmids, designated pSCS14-17, could be distinguished. All these plasmids conferred Cmr by encoding the Cm-inactivating enzyme chloramphenicol acetyltransferase (CAT). In all four types of Cmr plasmids from S. lentus, the expression of the cat gene was inducible with Cm, as demonstrated by enzymatic assay and polyacrylamide gel electrophoresis.
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PMID:Emerging chloramphenicol resistance in Staphylococcus lentus from mink following chloramphenicol treatment: characterisation of the resistance genes. 780 25

Several lines of evidence demonstrate that the DNA of the iridovirus frog virus 3 (FV3) is methylated in all 5'-CG-3' sequences both in virion DNA and in the intracellular viral DNA at late times after infection. The 5-methyldeoxycytidine residues in this viral DNA occur exclusively in 5'-CG-3' dinucleotide positions. We have cloned and determined the nucleotide sequence of the L1140 gene and its promoter from FV3 DNA. The gene encodes a 40-kDa protein. The results of transcriptional pattern analyses for this gene in fathead minnow fish cells document that this gene is transcribed exclusively late after FV3 infection. The L1140 gene and its promoter are fully methylated at late times after infection. We have been interested in resolving the apparent paradox that the methylated L1140 promoter is methylated and active late in FV3-infected cells. Of course, the possibility cannot be excluded that one or a few 5'-CG-3' sequences outside restriction endonuclease sites escaped de novo methylation after FV3 DNA replication. We have devised a construct that places the chloramphenicol acetyltransferase gene under the control of the L1140 promoter. Upon transfection, this construct exhibits activity only in FV3-infected BHK-21 hamster cells, not in uninfected BHK-21 cells. The fully 5'-CG-3' or 5'-GCGC-3' (HhaI) methylated, HpaII-mock-methylated, or unmethylated L1140 promoter-chloramphenicol acetyltransferase gene construct is active in FV3-infected BHK-21 cells, whereas the same construct 5'-CCGG-3' (HpaII) methylated has lost activity. Apparently, complete methylation of the late L1140 promoter in FV3 DNA is compatible with activity. However, a very specific 5'-CCGG-3' methylation pattern that does not naturally occur in authentic FV3 DNA in infected cells abrogates promoter function. These results further support the notion that very specific patterns of methylation are required to inhibit or inactivate viral promoters.
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PMID:A fully 5'-CG-3' but not a 5'-CCGG-3' methylated late frog virus 3 promoter retains activity. 788 71

Double-strand breaks introduced into DNA in vivo have been shown to enhance homologous recombination in a variety of chromosomal and extrachromosomal loci in Saccharomyces cerevisiae. To introduce double-strand breaks in DNA at defined locations in mammalian cells, we have constructed a mammalian expression vector for a modified form of I-Sce I, a yeast mitochondrial intron-encoded endonuclease with an 18-bp recognition sequence. Expression of the modified I-Sce I endonuclease in COS1 cells results in cleavage of model recombination substrates and enhanced extrachromosomal recombination, as assayed by chloramphenicol acetyltransferase activity and Southern blot analysis. Constitutive expression of the endonuclease in mouse 3T3 cells is not lethal, possibly due to either the lack of I-Sce I sites in the genome or sufficient repair of them. Expression of an endonuclease with such a long recognition sequence will provide a powerful approach to studying a number of molecular processes in mammalian cells, including homologous recombination.
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PMID:Expression of a site-specific endonuclease stimulates homologous recombination in mammalian cells. 801 16

To define the mechanisms of 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced transcription of the ornithine decarboxylase (ODC) gene, we isolated a genomic clone (hODC41B) of ODC from a human leukocyte genomic DNA library. The restriction endonuclease map, in comparison with the previously published sequences of the human ODC gene, indicated that hODC41B contained a 15.7-kb sequence that extended from the sixth exon to about 10 kb upstream of the ODC gene. A 2.5-kb genomic fragment containing the 5' flanking region and the first exon was subcloned and sequenced. Sequence analysis revealed multiple putative promoter/enhancer elements (a TATA box, a CAAT box, 17 GC boxes, and a cAMP-responsive element) but no consensus AP-1 sequences (TGAGTCA) in the 2.5-kb 5' flanking region. However, three AP-1 sequences were located in introns 3, 5, and 11. We constructed a series of chimeric genes containing part of the first exon and increasingly longer 5' flanking sequences of the ODC gene fused to either bacterial chloramphenicol acetyltransferase (CAT) or luciferase reporter genes. TPA inducibility was determined by transient transfection and measurement of CAT or luciferase expression in HeLa cells. The induction of CAT activity by TPA decreased with decreasing lengths of the 5' flanking sequences up to nt -82. The TPA induction from the construct -72 ODC CAT was threefold to sevenfold, and the TPA inducibility of the same fragment was about ninefold to 30-fold with the luciferase reporter gene. Further deletion analysis revealed TPA-responsive sequences in ODC nt -42 to +54. Gel mobility shift assays using alpha-32P-end labeled ODC nt -42 to +60 revealed that nt -42 to +60 specifically bound HeLa cell nuclear proteins. HeLa cell nuclear protein binding to ODC nt -42 to +60 could not be completely competed by AP-1-, AP-2-, AP-3-, or SP1-responsive sequences.
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PMID:Non-AP-1 tumor promoter 12-O-tetradecanoylphorbol-13-acetate-responsive sequences in the human ornithine decarboxylase gene. 804 98

Plasmids of 70MDa from 6 Shigella sonnei isolates originating in Kaohsiung, Taiwan were found. All of them encoded resistance to chloramphenicol and tetracycline. These plasmids belonged to the H II incompatibility group. The fertility inhibition property of plasmids was the fi- type. Restriction endonuclease analysis revealed the six plasmids all had identical restriction patterns with BamH I, Hind III and Nru I. All six plasmids carried the type I chloramphenicol acetyltransferase gene and the class B tetracycline resistance gene as determined by Southern hybridization with DNA probes of various antimicrobial resistance genes. The results demonstrated that S. sonnei isolates harboured the same R plasmid.
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PMID:Characterization of R plasmids from Shigella sonnei in Taiwan. 804 80

The chloramphenicol resistance gene (pp-cat) was cloned from a transferable R plasmid of Pasteurella piscicida, pSP9351, and the sequence of the gene was determined. Subcloning and deletion analysis localized the resistance gene, pp-cat, to within a 2.3 kb HincII-BamHI fragment. The fragment as a probe hybridized with the type I chloramphenicol acetyltransferase (CAT) gene and did not hybridize to CAT types II, III, and CAT-VA. The fragment hybridized to transferable R plasmids encoded with resistance to chloramphenicol, which were detected from P. piscicida isolated in different years. Nucleotide sequences of the coding and flanking regions of pp-cat (2031 bp) identified an open reading frame coding type I CAT of a molecular mass of about 25,000 Da. Comparison analysis of the sequences outside the cat open reading frame showed also that pp-cat has homology, in part, with the gene that coding for the endonuclease EcoRII and those that flank the cat gene derived from the Acinetobacter baumannii chromosome.
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PMID:The structure of the chloramphenicol resistance gene on a transferable R plasmid from the fish pathogen, Pasteurella piscicida. 827 73

We have isolated and characterised the 5' region of a member of the carp myosin heavy chain gene family. Expression of this gene has previously been shown to be induced by an increase in environmental temperature and is restricted to the small-diameter white myotomal muscle fibres which are associated with growth. The whole isoform gene, including potential regulatory sequence 5' to the transcription start site and the 3' untranslated region was cloned in a lambda2001 bacteriophage vector. Studies of the structure of the 5'-end of the gene revealed high amino acid sequence similarity with translated exons 3-7 of mammalian myosin heavy chain genes indicating identical exon/intron boundaries. The overall length of the gene was however only about one half of that in mammals and birds due to shorter introns. The region 5' to the transcription unit was sequenced and revealed the presence of putative TATA and CCAAAT boxes. In order to study the regulation of expression, a series of endonuclease-generated fragments from the 5' flanking sequence were spliced to chloramphenicol acetyltransferase reporter vectors and used in cell transfection assays or direct gene injection into carp skeletal muscle. The 5' flanking region, which contains a consensus sequence known as an E-box (CANNTG) and a MEF2 binding site, was shown to improve the expression of the reporter gene in fish acclimated at 18 degrees C or 28 degrees C. Unlike the coding region, there was little similarity between the 5'-upstream sequence (promoter region) when compared with sequences flanking the 5'-end of the other myosin heavy chain genes in mammals or chicken.
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PMID:The characterisation of the 5' regulatory region of a temperature-induced myosin-heavy-chain gene associated with myotomal muscle growth in the carp. 866 10

The use of viral vectors to deliver foreign genes offers some promise of generating new and more efficacious vaccines. However, the insertion of foreign genes into viral genomes often results in the insertional mutagenesis of one or more genes that adversely affect replication. In an attempt to overcome this problem, we constructed two portable intron cassettes. The cassettes were derived from the adenovirus late leader 1 intron and were cloned into either the chloramphenicol acetyltransferase (CAT) gene or the LacZ gene of Escherichia coli. The intron cassettes were transfected into chicken embryo fibroblasts (CEFs) and the cell lysates were later assayed for either beta-galactosidase (beta-Gal) or CAT activity. The first intron cassette (type A) contained flanking adenovirus exon sequences. Consequently, the flanking adenovirus exon sequences remained in the spliced transcript. With the type A intron inserted in the correct orientation for splicing, CAT activity was not diminished. However, in the reverse orientation, no CAT activity could be detected. The second intron cassette (type B) had the splice donor and splice acceptor sites converted to the blunt-end restriction endonuclease sites Pml I and Pvu II, respectively. The blunt-end restriction endonuclease sites enabled the portable intron to be removed from the flanking adenovirus exon sequences and inserted into any blunt-end restriction endonuclease site in the recipient gene. After splicing, no adenovirus exon sequences remained in the recipient gene's RNA transcript. To demonstrate its usefulness, an insertion cassette was made by cloning the E. coli LacZ gene into a multiple cloning site within the type B intron. The insertion cassette was then cloned into a Pvu II site in the middle of the CAT gene. Following transfection in CEFs, high levels of both CAT and beta-Gal were detected, demonstrating that both genes were properly transcribed and translated.
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PMID:Construction of portable intron cassettes for the delivery and expression of foreign genes. 898 25

We investigated the minimal promoter of APEX, which encodes mouse apurinic DNA repair endonuclease. A 1.85-kb fragment with APEX upstream sequences and approximately 290 bp of the transcribed region linked to a chloramphenicol acetyltransferase (CAT) reporter gene was assayed by transient transfection in NIH-3T3 cells. The minimal APEX promoter was comprised of approximately 190 bp of upstream and approximately 170 bp of transcribed DNA (exon 1 and most of intron 1). This approximately 360-bp region contains two CCAAT boxes and other consensus protein binding sites, but no TATA box. Deletion of the 5'-most CCAAT box decreased activity approximately 5-fold. The second CCAAT box (situated in exon 1) may play an independent role in APEX expression. Transcription start sites have been identified downstream of the second CCAAT box, and DNase I footprinting demonstrated NIH-3T3 nuclear proteins binding this region, including an Spl site located between the CCAAT boxes. Electrophoretic mobility-shift assays indicated binding by purified Sp1. Mouse proteins did not bind three myc-like (USF) sites in the APEX promoter, in contrast to the APE promoter. The APEX and APE promoter had similar activity in Hela cells, but in mouse cells, the murine promoter had approximately 5-fold higher activity than did the human promoter. Both the APEX and APE promoters exhibited bidirectional activity in their cognate cells.
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PMID:Comparison of the promoters of the mouse (APEX) and human (APE) apurinic endonuclease genes. 950 86


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