Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.7.48 (transcriptase)
 9,479 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In plants, fatty acids synthesized in the chloroplasts are exported as acyl-CoA esters to the endoplasmic reticulum (ER). Cytosolic 10-kDa acyl-CoA-binding proteins (ACBPs), prevalent in eukaryotes, are involved in the storage and intracellular transport of acyl-CoAs. We have previously characterized Arabidopsis thaliana cDNAs encoding membrane-associated ACBPs with ankyrin repeats, designated ACBP1 and ACBP2, which show conservation to cytosolic ACBPs at the acyl-CoA-binding domain. Analysis of the Arabidopsis genome has revealed the presence of three more genes encoding putative proteins with acyl-CoA-binding domains, designated ACBP3, ACBP4 and ACBP5. Homologues of ACBP1 to ACBP5 have not been reported in any other organism. We show by reverse-transcriptase polymerase chain reaction (RT-PCR) analysis that ACBP3 , ACBP4 and ACBP5 are expressed in all plant organs, like ACBP1 and ACBP2 . ACBP4 and ACBP5 that share 81.4 identity and which contain kelch motifs were further investigated. To demonstrate their function in binding acyl-CoA, we have expressed them as (His)6-tagged recombinant proteins in Escherichia coli for in vitro binding assays. Both (His)6-ACBP4 and (His)6-ACBP5 bind [14C]oleoyl-CoA with high affinity, [14C]palmitoyl-CoA with lower affinity and did not bind [14C]arachidonyl-CoA. Eight mutant forms of each protein with single amino acid substitutions within the acyl-CoA-binding domain were produced and analyzed. On binding assays, all mutants were impaired in oleoyl-CoA binding. Hence, these novel ACBPs with kelch motifs have functional acyl-CoA-binding domains that bind oleoyl-CoA. Their predicted cytosol localization suggests that they could maintain an oleoyl-CoA pool in the cytosol or transport oleoyl-CoA from the plastids to the ER in plant lipid metabolism.
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PMID:ACBP4 and ACBP5, novel Arabidopsis acyl-CoA-binding proteins with kelch motifs that bind oleoyl-CoA. 1560 82

Arabidopsis thaliana acyl-CoA-binding protein 2 (ACBP2) was observed to interact with farnesylated protein 6 (AtFP6), which has a metal-binding motif (M/LXCXXC). Their interaction and expression in response to heavy metals were investigated. Yeast two-hybrid analysis and in vitro assays showed that an ACBP2 derivative lacking ankyrin repeats did not interact with AtFP6, indicating that the ankyrin repeats mediate protein-protein interaction. Autofluorescence-tagged ACBP2 and AtFP6 transiently co-expressed in tobacco (Nicotiana tabacum) were both targeted to the plasma membrane. Reverse transcriptase polymerase chain reaction and northern blot analyses revealed that AtFP6 mRNA was induced by cadmium (Cd(II)) in A. thaliana roots. Assays using metal-chelate affinity chromatography demonstrated that in vitro translated ACBP2 and AtFP6 bound lead (Pb(II)), Cd(II) and copper (Cu(II)). Consistently, assays using fluorescence analysis confirmed that (His)(6)-AtFP6 bound Pb(II), like (His)(6)-ACBP2. Arabidopsis thaliana plants overexpressing ACBP2 or AtFP6 were more tolerant to Cd(II) than wild-type plants. Plasma membrane-localized ACBP2 and AtFP6 probably mediate Pb(II), Cd(II) and Cu(II) transport in A. thaliana roots. Also, (His)(6)-ACBP2 binds [(14)C]linoleoyl-CoA and [(14)C]linolenoyl-CoA, the precursors for phospholipid repair following lipid peroxidation under heavy metal stress at the plasma membrane. ACBP2-overexpressing plants were more tolerant to hydrogen peroxide than wild-type plants, further supporting a role for ACBP2 in post-stress membrane repair.
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PMID:Arabidopsis thaliana acyl-CoA-binding protein ACBP2 interacts with heavy-metal-binding farnesylated protein AtFP6. 1882 12

In the classic retinoid cycle, 11-cis retinol is synthesized in the retinal pigment epithelium (RPE) by two enzymes: Isomerase I (RPE65) and lecithin:retinol acyltransferase (LRAT). The purpose of this study is to provide experimental evidence for two active isomerases in the cone-dominated chicken eye: an LRAT-dependent Isomerase I in the RPE and an ARAT (acyl CoA:retinol acyltransferase)-dependent isomerase (Isomerase II) in the retina. First, we show that whole chicken retina in vitro, removed from the RPE/choroid and sclera, produces 11-cis retinoids upon light exposure, indicating the existence of RPE-independent isomerase (Isomerase II) activity in the retina. Reverse transcriptase polymerase chain reaction studies show high levels of RPE65 expression in the RPE, low levels in the retina, and none in primary Muller cell cultures, indicating the presence of Isomerase I in the RPE and a minimal amount in the retina. Activities of the RPE and retina isomerases were then measured by enzyme assays with specific enzyme inhibitors. 2,2'-Bipyridine, a known Isomerase I inhibitor, and N-ethylmaleimide (NEM), a known LRAT inhibitor, significantly reduced Isomerase I activity but not Isomerase II activity. Progesterone, a known ARAT inhibitor, completely blocked Isomerase II activity but not Isomerase I activity. Thus, this study reports novel results for distinguishing the biochemical properties of Isomerase I from those of Isomerase II, as well a difference in their locations in the chicken eye. On the basis of these differences, the cone-dominated chicken eye must contain two retinoid cycles: a classic visual cycle for retinoid exchange between the RPE and the retina supported by Isomerase I in the RPE and an additional visual cycle for retinoid processing in the retina supported by Isomerase II.
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PMID:Evidence for two retinoid cycles in the cone-dominated chicken eye. 1949 94

The PA, PB1, and PB2 subunits, components of the RNA-dependent RNA polymerase of influenza A virus, are essential for viral transcription and replication. The PB2 subunit binds to the host RNA cap (7-methylguanosine triphosphate (m(7)GTP)) and supports the endonuclease activity of PA to "snatch" the cap from host pre-mRNAs. However, the structure of PB2 is not fully understood, and the functional sites remain unknown. In this study, we describe a novel Val/Arg/Gly (VRG) site in the PB2 cap-binding domain, which is involved in interaction with acetyl-CoA found in eukaryotic histone acetyltransferases (HATs). In vitro experiments revealed that the recombinant PB2 cap-binding domain that includes the VRG site interacts with acetyl-CoA; moreover, it was found that this interaction could be blocked by CoA and various HAT inhibitors. Interestingly, m(7)GTP also inhibited this interaction, suggesting that the same active pocket is capable of interacting with acetyl-CoA and m(7)GTP. To elucidate the importance of the VRG site on PB2 function and viral replication, we constructed a PB2 recombinant protein and recombinant viruses including several patterns of amino acid mutations in the VRG site. Substitutions of the valine and arginine residues or of all 3 residues of the VRG site to alanine significantly reduced the binding ability of PB2 to acetyl-CoA and its RNA polymerase activity. Recombinant viruses containing the same mutations could not be replicated in cultured cells. These results indicate that the PB2 VRG sequence is a functional site that is essential for acetyl-CoA interaction, RNA polymerase activity, and viral replication.
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PMID:A novel functional site in the PB2 subunit of influenza A virus essential for acetyl-CoA interaction, RNA polymerase activity, and viral replication. 2506 5

The PA, PB1, and PB2 subunits, components of the RNA-dependent RNA polymerase of influenza A virus, and the nucleoprotein (NP) interact with the genomic RNA of influenza viruses and form ribonucleoproteins. Especially, the PB2 subunit binds to the host RNA cap [7-methylguanosine triphosphate (m7GTP)] and supports the endonuclease activity of PA to "snatch" the cap from host pre-mRNAs. In this study, we describe a novel Val/Arg/Gly (VRG) site in the PB2 cap-binding domain, which is necessary for interaction with acetyl-CoA found in eukaryotic histone acetyltransferases (HATs). In vitro experiments revealed that the recombinant PB2 cap-binding domain that includes the VRG site interacts with acetyl-CoA; moreover, it was found that this interaction could be blocked by CoA and various HAT inhibitors. Interestingly, m7GTP also inhibited this interaction, suggesting that the same active pocket is capable of interacting with acetyl-CoA and m7GTP. To elucidate the importance of the VRG site on PB2 function and viral replication, we constructed a PB2 recombinant protein and recombinant viruses including several patterns of amino acid mutations in the VRG site. Substitutions of 2 or 3 amino acid residues of the VRG site to alanine significantly reduced PB2's binding ability to acetyl-CoA and its RNA polymerase activity. Recombinant viruses containing the same mutations could not be replicated in cultured cells. These results indicate that the PB2 VRG sequence is a functional site that is essential for acetyl-CoA interaction, RNA polymerase activity, and viral replication. I will also discuss some novel functions of NP in this review.
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PMID:Structural and Biochemical Analyses on the RNA-dependent RNA Polymerase of Influenza Virus for Development of Novel Anti-influenza Agents. 2815 33