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)

A full length human androgen receptor (hAR) cDNA was constructed from cDNA and genomic clones. Structurally the 10.6-kilobase (kb) hAR cDNA consists of a long 5'-untranslated region (5'-UTR, 1.1 kb), a previously described open reading frame (ORF, 2.7 kb) (Trapman, J., Klaassen, P., Kuiper, G. G. J. M., van der Korput, J. A. G. M., Faber, P. W., van Rooij, H. C. J., Geurts van Kessel, A., Voorhorst, M. M., Mulder, E., and Brinkmann, A. O. (1988) Biochem. Biophys. Res. Commun. 153, 241-248; Faber, P. W., Kuiper, G. G. J. M., van Rooij, H. C. J., van der Korput, J. A. G. M., Brinkmann, A. O., and Trapman, J. (1989) Mol. Cell. Endocrinol. 61, 257-262), and a very long 3'-untranslated region (3'-UTR, 6.8 kb). The complete 5'- and 3'-UTRs were found to be encoded by the previously reported first and eight protein coding exons of the hAR gene, respectively (Kuiper, G. G. J. M., Faber, P. W., van Rooij, H. C. J., van der Korput, J. A. G. M., Ris-Stalpers, C., Klaassen, P., Trapman, J., and Brinkmann, A. O. (1989) J. Mol. Endocrinol. 2, R1-R4). Two major sites of transcription initiation were identified in a 13-base pair region. DNA fragments spanning these transcription initiation sites conferred promoter activity upon a promoterless chloramphenicol acetyltransferase reporter gene construct. Two equally effective, functional polyadenylation signals (ATTAAA and CATAAA) at a mutual distance of 221 base pairs were detected. The ATTAAA hexamer sequence gave rise to multiple sites of poly(A) addition, whereas only one position was used following the CATAAA hexamer. In LNCaP prostatic carcinoma cells an alternatively spliced hAR mRNA species was identified which lacks 3 kb of the 3'-UTR.
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PMID:Characterization of the human androgen receptor transcription unit. 171 Feb 13

We have mapped a transcriptional regulatory sequence within the 1.0 kb noncoding region of the bovine papilloma virus (BPV-1) genome, using an enhancer dependent expression vector for chloramphenicol acetyltransferase. This transcriptional regulatory element works independently of position and orientation, and its function is significantly augmented in BPV-1 transformed C127 cells and in monkey CV-1 cells acutely transfected with plasmids expressing BPV-1 early gene products. Using defined deletion mutants of the BPV-1 DNA and full-length viral cDNAs expressed from an SV40 early promoter, we demonstrate that the expression of this trans-activating factor maps to the 3' open reading frames of the viral transforming region. A premature termination codon engineered into the E2 ORF eliminates expression of this diffusible transactivation function establishing the E2 gene product as the diffusible trans-activating factor.
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PMID:Transactivation of a bovine papilloma virus transcriptional regulatory element by the E2 gene product. 299 Jul 24

Promoter elements responsible for directing the transcription of six tightly clustered vaccinia virus (VV) late genes (open reading frames [ORFs] D11, D12, D13, A1, A2, and A3) from the HindIII D/A region of the viral genome were identified within the upstream sequences proximal to each individual locus. These regions were identified as promoters by excising them from the VV genome, abutting them to the bacterial chloramphenicol acetyl transferase gene, and demonstrating their ability to drive expression of the reporter gene in transient-expression assays in an orientation-specific manner. To delineate the 5' boundary of the upstream elements, two of the VV late gene (A1 and D13) promoter: CAT constructs were subjected to deletion mutagenesis procedures. A series of 5' deletions of the ORF A1 promoter from -114 to -24 showed no reduction in promoter activity, whereas additional deletion of the sequences from -24 to +2 resulted in the complete loss of activity. Deletion of the ORF A1 fragment from -114 to -104 resulted in a 24% increase in activity, suggesting the presence of a negative regulatory region. In marked contrast to previous 5' deletion analyses which have identified VV late promoters as 20- to 30-base-pair cap-proximal sequences, 5' deletions to define the upstream boundary of the ORF D13 promoter identified two positive regulatory regions, the first between -235 and -170 and the second between -123 and -106. Background levels of chloramphenicol acetyltransferase expression were obtained with deletions past -88. Significantly, this places the ORF D13 regulatory regions within the upstream coding sequences of the ORF A1. A high-stringency computer search for homologies between VV late promoters that have been thus far characterized was carried out. Several potential consensus sequences were found just upstream from RNA start sites of temporally related promoter elements. Three major conclusions are drawn from these experiments. (i) The presence of promoters preceding each late ORF supports the hypothesis that each is expressed as an individual transcriptional unit. (ii) Promoter elements can be located within the coding portion of the upstream gene. (iii) Sequence homologies between temporally related promoter elements support the notion of kinetic subclasses of late genes.
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PMID:Molecular dissection of cis-acting regulatory elements from 5'-proximal regions of a vaccinia virus late gene cluster. 333 46

The UL3 protein of equine herpesvirus 1 (EHV-1) KyA strain is a homolog of the ICP27 alpha regulatory protein of herpes simplex virus type 1 (HSV-1) and the ORF 4 protein of varicella-zoster virus. To characterize the regulatory function of the UL3 gene product, a UL3 gene expression vector (pSVUL3) and a vector expressing a truncated version of the UL3 gene (pSVUL3P) were generated. These effector plasmids, in combination with an EHV-1 immediate-early (IE) gene expression vector (pSVIE) and chimeric EHV-1 promoter-chloramphenicol acetyltransferase (CAT) reporter constructs, were used in transient transfection assays. These assays demonstrated that the EHV-1 UL3 gene product is a regulatory protein that can independently trans activate the EHV-1 IE promoter; however, this effect can be inhibited by the repressive function of the IE gene product on the IE promoter (R. H. Smith, G. B. Caughman, and D. J. O'Callaghan, J. Virol. 66:936-945, 1992). In the presence of the IE gene product, the UL3 gene product significantly augments gene expression directed by the promoters of three EHV-1 early genes (thymidine kinase; IR4, which is the homolog of HSV-1 ICP22; and UL3 [ICP27]) and the promoter of the EHV-1 late gene IR5, which is the homolog of HSV-1 US10. Sequences located at nucleotides -123 to +20 of the UL3 promoter harbor a TATA box, SP1 binding site, CAAT box, and octamer binding site and, when linked to the CAT reporter gene, are trans activated to maximal levels by the pSVIE construct in transient expression assays. Results from CAT assays also suggest that the first 11 amino acids of the UL3 protein are not essential for the regulatory function of the UL3 gene product.
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PMID:Regulatory function of the equine herpesvirus 1 ICP27 gene product. 770

We have made use of certain novel genetic elements of picornaviruses termed internal ribosomal entry sites (IRES) to construct a viral RNA with the following genetic order: PV 5' NTR-EMCV IRES-PV ORF-3' NTR (PV, poliovirus; NTR, nontranslated region; EMCV, encephalomyocarditis virus; ORF, open reading frame). Transfection of this RNA into HeLa cells yielded a poliovirus (W1-PNENPO) that contained two heterologous IRES elements (type 1 IRES of PV; type 2 IRES of EMCV) in tandem. The insertion of foreign coding sequences into the genome of W1-PNENPO between the IRES elements yielded viable polioviruses with the gene order PV 5' NTR-foreign ORF-EMCV IRES-PV ORF-3' NTR. The foreign ORFs we have employed in this study included the coding region for chloramphenicol acetyltransferase (CAT), or segments of either luciferase or the HIV-1 envelope glycoprotein gp120. W1-PV/V3-3, a dicistronic poliovirus that contained HIV-1-specific sequences that included the V3 domain of gp120, was used to infect transgenic mice (PVR+) that were engineered to express the poliovirus receptor. The genetic stability of the dicistronic viruses and the HIV-1-specific immune response in PVR+ mice after infection with these novel agents are discussed.
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PMID:Dicistronic polioviruses as expression vectors for foreign genes. 786 34

The primary 2A/2B polyprotein cleavage of aphtho-and cardioviruses is mediated by their 2A proteins cleaving C-terminally. Whilst the aphthovirus 2A region is only 16 aa (possibly 18 aa) long, the cardiovirus 2A protein is some 150 aa. We have previously shown that foot-and-mouth disease virus (FMDV) 2A is able to mediate cleavage in an artificial (chloramphenicol acetyltransferase/FMDV 2A/beta-glucuronidase [CAT-2A-GUS]) polyprotein system devoid of any other FMDV sequences with high (approximately 85%), although not complete, cleavage. In this paper we show that insertion of upstream FMDV capsid protein 1 D sequences increases the activity. In addition, we have demonstrated that the cardiovirus Theiler's murine encephalomyelitis virus(TME) 2A protein, when linked to GUS in a single ORF, is able to cleave at its own C terminus with high efficiency--if not completely. The C-terminal 19 aa of TME 2A, together with the N-terminal proline residue of protein 2B, were inserted into the CAT/GUS artificial polyprotein system (in a single ORF). This recombinant [CAT-deltaTME2A-GUS] polyprotein was able to mediate cleavage with high (approximately 85%) efficiency--directly comparable to the activity observed when FMDV 2A was inserted. A similar insertion into [CAT-GUS] of the C-terminal 19 aa of the cardiovirus encephalomyocarditis virus (EMC) 2A, together with the N-terminal proline residue of protein 2B, produced a [CAT-delta EMC2A-GUS] polyprotein which also mediated cleavage at approximately 85%. Analysis of the products of expression of these artificial polyproteins in a prokaryotic translation system did not, apparently, reveal any GUS cleavage product.
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PMID:The cleavage activities of aphthovirus and cardiovirus 2A proteins. 901 Feb 80

A new insertion sequence element, IS1452, was found to be associated with inactivation of the alcohol dehydrogenase by insertion in the adhS gene encoding subunit III of the three-component membrane-bound alcohol dehydrogenase complex in Acetobacter pasteurianus. Cloning and sequencing analyses of the mutated subunit III gene locus revealed that IS1452 was inserted at or near the ribosome-binding sequence of adhS. Analysis of transcription using the chloramphenicol acetyltransferase gene as the reporter indicated that IS1452 abolished transcription of adhS by separating its promoter from the subunit III structural gene. IS1452 was 1411 bp in length and had a terminal inverted repeat of 21 bp. IS1452 contained one long ORF of 416 amino acids rich in basic amino acids. This protein showed homology with a putative transposes, Tra1, of IS701 isolated from the cyanobacterium Calothrix species PCC 7601. Like IS701, IS1452 was found to generate a 4 bp direct repeat at the site of insertion upon transposition. The target site specificity was rather strict, and a CTA(A or G) sequence appeared to be preferentially recognized. Transposition of IS1452 was replicative, since it was accompanied by an increase in the copy number of IS1452. Several strains belonging to the genus Acetobacter also contained IS1452 at varying copy numbers from one to more than ten. These observations suggest that IS1452 is one of the insertion sequences that are responsible for genetic instability leading to deficiencies in various physiological properties in acetic acid bacteria.
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PMID:A new insertion sequence IS1452 from Acetobacter pasteurianus. 904 30

The gene encoding acetyl CoA:deacetylvindoline 4-O-acetyltransferase (DAT) (EC 2.3.1.107) which catalyzes the last step in vindoline biosynthesis was isolated and characterized. The genomic clone encoded a 50 kDa polypeptide containing the sequences of nine tryptic fragments derived from the purified DAT heterodimer. However, cleavage of DAT protein to yield a heterodimer appears to be an artifact of the protein purification procedure, since the size of the protein (50 kDa) cross-reacting with anti-DAT antibody in seedlings and in leaves of various ages also corresponds to the size of the active recombinant enzyme. Studies with the intact plant and with developing seedlings showed that induction of DAT mRNA, protein accumulation and enzyme activity occurred preferentially in vindoline producing tissues such as leaves and cotyledons of light-treated etiolated seedlings. The ORF of DAT showed significant sequence identity to 19 other plant genes, whose biochemical functions were mostly unknown. The Mr of approximately 50 kDa, a HXXXDG triad, and a DFGWGKP consensus sequence are highly conserved among the 20 plant genes and these criteria may be useful to identify this type of acyltransferase. The involvement of some of these genes in epicuticular wax biosynthesis, fruit-ripening and in benzoyltransfer reactions indicates that the plant kingdom contains a superfamily of multifunctional acyltransferases which operate by a reaction mechanism related to the ancient chloramphenicol O-acetyltransferase and dihydrolipoyl acetyltransferase class of enzymes.
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PMID:The terminal O-acetyltransferase involved in vindoline biosynthesis defines a new class of proteins responsible for coenzyme A-dependent acyl transfer. 968 Oct 34

Protein synthesis directed by hepatitis A virus (HAV) RNA is mediated by a mechanism involving the recognition of internal sequences. Two in-frame AUG codons initiate the long open reading frame (positions 734-736 and 740-742). The extra-cistronic region extending between the uncapped 5'-end and the ORF contains two pyrimidine-rich tracts (PRTs): one 12 nucleotides in length in the close vicinity of the initiator AUG, and a longer one between bases 94 and 140. In order to study the relative contribution of these elements to the process of internal initiation of translation, cDNA representations of the 5'-terminal extra-cistronic region of HAV RNA were inserted in the intergenic region of the bi-cistronic plasmid pSV-GH/CAT, between the genes encoding the human growth hormone (GH) and the bacterial enzyme chloramphenicol acetyltransferase (CAT), and following transfection of COS-1 cells, the transient expression of both genes was quantified. The importance of the 3'-PRT appeared to be strongly influenced by the length of the 'spacer' sequence extending between this structure and the translation initiation site: placed 45 nucleotides upstream from the initiator codon of a reporter gene, its integrity was stringently required for initiation to occur. Bringing the length of the 'spacer' back to its actual size in HAV RNA (i.e. 11 or 17 nt) reduced considerably the overall rate of internal initiation of translation, and the relative contribution to this process of the 3'-PRT became marginal. Concomitantly, the importance of the functional domains previously identified in the 5'-PRT fluctuated: while integrity of domain 100-106 was always stringently required for initiation to occur, the activity of domain 113-118 paralleled that of the 3'-PRT, and the opposite applied to domain 121-126, whose contribution became relevant only after switching off the 3'-PRT. Systematic mutations introduced in the 'spacer' sequences suggest that the length of this region may be responsible for the down regulation of translation of HAV RNA and, possibly, for its lengthy replication cycle.
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PMID:The distance between the 3'-pyrimidine-rich tract and the AUG codon modulates internal initiation of translation of hepatitis A virus RNA. 973 41

Translation of the open reading frame 2 (ORF-2) of the human respiratory syncytial virus M2 gene initiates at one of the three initiation codons located upstream of the termination codon for the first ORF. Replacement of ORF-2 with the major ORF of the chloramphenicol acetyltransferase reporter gene followed by systematic mutagenesis of the putative initiation codons demonstrated the usage of these codons as the translational initiators for ORF-2 expression both in vitro and in vivo. While the efficiency of translation was maintained when only the first and second AUG codons were preserved in vivo, there was no apparent preference in vitro for any of the three codons when only one was present. Mutagenesis studies showed that the location of the termination codon of ORF-1 protein plays a crucial role in directing translation of ORF-2 from the upstream initiation codons in vivo. This indicates that the second ORF is accessed by the ribosomes that are departing from the first ORF and that these ribosomes reinitiate on AUG codons 5' to the point of translation termination.
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PMID:Expression of the ORF-2 protein of the human respiratory syncytial virus M2 gene is initiated by a ribosomal termination-dependent reinitiation mechanism. 1083 65


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