Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.6 (RNA polymerase)
 34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We describe a method for specifically labelling T7 RNA polymerase at (or near) the active site. Enzyme molecules that have been modified by covalent attachment of a benzaldehyde nucleotide derivative in the presence of template DNA are subsequently incubated with radioactively labelled nucleoside triphosphates. Labelling of the enzyme occurs as a result of the formation of the first phosphodiester bond. The labelling is template-directed and the expected specificity of initiation at individual T7 promoters is observed. The label has been localized to an 80 kd tryptic fragment that contains the carboxy-terminal portion of the enzyme.
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PMID:Specific labelling of the active site of T7 RNA polymerase. 368 73

DNA-dependent RNA polymerase B (II) from wheat germ was modified by incubation with 4-[N-(beta-hydroxyethyl)-N-methyl]benzaldehyde esters of AMP, ADP or ATP, followed by reduction with NaBH4. Reaction of the modified enzyme with [alpha-32P]UTP in the presence of various DNA templates led to a highly selective affinity labelling of the subunit with Mr 140 000 by covalently linked ApU. Labelling was inhibited by 1 microgram/ml alpha-amanitin.
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PMID:Highly selective affinity labelling of RNA polymerase B (II) from wheat germ. 370 95

Various cinnammoyl-based structures were synthesized and tested in enzyme assays as inhibitors of the HIV-1 integrase (IN). The majority of compounds were designed as geometrically or conformationally constrained analogues of caffeic acid phenethyl ester (CAPE) and were characterized by a syn disposition of the carbonyl group with respect to the vinylic double bond. Since the cinnamoyl moiety present in flavones such as quercetin (inactive on HIV-1-infected cells) is frozen in an anti arrangement, it was hoped that fixing our compounds in a syn disposition could favor anti-HIV-1 activity in cell-based assays. Geometrical and conformational properties of the designed compounds were taken into account through analysis of X-ray structures available from the Cambridge Structural Database. The polyhydroxylated analogues were prepared by reacting 3,4-bis(tetrahydropyran-2-yloxy)benzaldehyde with various compounds having active methylene groups such as 2-propanone, cyclopentanone, cyclohexanone, 1,3-diacetylbenzene, 2, 4-dihydroxyacetophenone, 2,3-dihydro-1-indanone, 2,3-dihydro-1, 3-indandione, and others. While active against both 3'-processing and strand-transfer reactions, the new compounds, curcumin included, failed to inhibit the HIV-1 multiplication in acutely infected MT-4 cells. Nevertheless, they specifically inhibited the enzymatic reactions associated with IN, being totally inactive against other viral (HIV-1 reverse transcriptase) and cellular (RNA polymerase II) nucleic acid-processing enzymes. On the other hand, title compounds were endowed with remarkable antiproliferative activity, whose potency correlated neither with the presence of catechols (possible source of reactive quinones) nor with inhibition of topoisomerases. The SARs developed for our compounds led to novel findings concerning the molecular determinants of IN inhibitory activity within the class of cinnamoyl-based structures. We hypothesize that these compounds bind to IN featuring the cinnamoyl residue C=C-C=O in a syn disposition, differently from flavone derivatives characterized by an anti arrangement about the same fragment. Certain inhibitors, lacking one of the two pharmacophoric catechol hydroxyls, retain moderate potency thanks to nonpharmacophoric fragments (i.e., a m-methoxy group in curcumin) which favorably interact with an "accessory" region of IN. This region is supposed to be located adjacent to the binding site accommodating the pharmacophoric dihydroxycinnamoyl moiety. Disruption of coplanarity in the inhibitor structure abolishes activity owing to poor shape complementarity with the target or an exceedingly high strain energy of the coplanar conformation.
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PMID:Geometrically and conformationally restrained cinnamoyl compounds as inhibitors of HIV-1 integrase: synthesis, biological evaluation, and molecular modeling. 976 32

The areCBA genes in Acinetobacter sp. strain ADP1, determining growth on benzyl alkanoates, are shown to be transcribed as a single operon and regulated by areR, which encodes a regulatory protein of the NtrC/XylR family. Assays of the Are enzymes and of two insertions of lacZ as a reporter gene have shown that the operon is induced by benzyl acetate, benzyl alcohol, and benzaldehyde, as well as 2- and 4-hydroxybenzyl acetates and benzyl propionate and butyrate. Two adjacent sites of transcriptional initiation were 97 and 96 bp upstream of the start codon for areC, near a sigma(54)-dependent -12, -24 promoter. Inactivation of areR and rpoN (for RNA polymerase sigma(54)) drastically reduced growth rates on the Are substrates and induction of the operon.
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PMID:areCBA is an operon in Acinetobacter sp. strain ADP1 and Is controlled by AreR, a sigma(54)-dependent regulator. 1111 44

We have previously reported the in vitro selection of a ribozyme that catalyzes an aldol reaction between a levulinic amide aldol donor and a benzaldehyde substrate. The selection scheme involved the priming of the RNA library with a levulinic amide aldol donor group that was introduced via transcription priming in the presence of a modified guanosine mononucleotide derivative. Here we provide a detailed description of the synthesis of the ribozyme substrates and the substrate oligonucleotides used for its isolation and characterization. The aldol donor group was attached to the phosphate moiety of guanosine monophosphate via a photocleavable linker molecule. This initiator nucleotide was efficiently incorporated into RNA molecules of differing sizes and composition by transcription priming with T7 RNA polymerase. With this method modified RNA oligonucleotides as small as a 6-mer sequence can be generated. A temperature profile of the intermolecular reaction indicates that the modified RNA hexamer binds the ribozyme largely by Watson-Crick pairing and only to a minor extent via the non-RNA moiety, whereas the ribozyme appears to have evolved a specific binding site for the aldehyde substrate.
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PMID:Photocleavable initiator nucleotide substrates for an aldolase ribozyme. 1851 52