RESUMO
Post-transcriptional autoregulation of gene expression is common in bacteria but many fewer examples are known in eukaryotes. We used the yeast collection of genes fused to GFP as a rapid screen for examples of feedback regulation in ribosomal proteins by overexpressing a non-regulatable version of a gene and observing the effects on the expression of the GFP-fused version. We tested 95 ribosomal protein genes and found a wide continuum of effects, with 30% showing at least a 3-fold reduction in expression. Two genes, RPS22B and RPL1B, showed over a 10-fold repression. In both cases the cis-regulatory segment resides in the 5' UTR of the gene as shown by placing that segment of the mRNA upstream of GFP alone and demonstrating it is sufficient to cause repression of GFP when the protein is over-expressed. Further analyses showed that the intron in the 5' UTR of RPS22B is required for regulation, presumably because the protein inhibits splicing that is necessary for translation. The 5' UTR of RPL1B contains a sequence and structure motif that is conserved in the binding sites of Rpl1 orthologs from bacteria to mammals, and mutations within the motif eliminate repression.
Assuntos
Retroalimentação Fisiológica , Proteínas Fúngicas/genética , Regulação Fúngica da Expressão Gênica , Proteínas Ribossômicas/genética , Sequência de Bases , Proteínas Fúngicas/química , Proteínas Fúngicas/metabolismo , Ordem dos Genes , Genes Reporter , Homeostase , Plasmídeos/genética , Proteínas Ribossômicas/química , Proteínas Ribossômicas/metabolismoRESUMO
Methylation of CpG (cytosine-phosphate-guanine) dinucleotides is a common epigenetic mark that influences gene expression. The effects of methylation on transcription factor (TF) binding are unknown for most TFs and, even when known, such knowledge is often only qualitative. In reality, methylation sensitivity is a quantitative effect, just as changes to the DNA sequence have quantitative effects on TF binding affinity. We describe Methyl-Spec-seq, an easy-to-use method that measures the effects of CpG methylation (mCPG) on binding affinity for hundreds to thousands of variants in parallel, allowing one to quantitatively assess the effects at every position in a binding site. We demonstrate its use on several important DNA binding proteins. We calibrate the accuracy of Methyl-Spec-seq using a novel two-color competitive fluorescence anisotropy method that can accurately determine the relative affinities of two sequences in solution. We also present software that extends standard methods for representing, visualizing, and searching for matches to binding site motifs to include the effects of methylation. These tools facilitate the study of the consequences for gene regulation of epigenetic marks on DNA.
Assuntos
Metilação de DNA , DNA/metabolismo , Fatores de Transcrição/metabolismo , Animais , Fator de Ligação a CCCTC/química , Fator de Ligação a CCCTC/metabolismo , Ilhas de CpG , DNA/química , Polarização de Fluorescência , Proteínas de Homeodomínio/química , Proteínas de Homeodomínio/metabolismo , Camundongos , Receptor Patched-1/química , Receptor Patched-1/metabolismo , Ligação Proteica , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Proteínas Repressoras/química , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Fatores de Transcrição/químicaRESUMO
The quantitative specificity of the STAT1 transcription factor was determined by measuring the relative affinity to hundreds of variants of the consensus binding site including variations in the length of the site. The known consensus sequence is observed to have the highest affinity, with all variants decreasing binding affinity considerably. There is very little loss of binding affinity when the CpG within the consensus binding site is methylated. Additionally, the specificity of mutant proteins, with variants of amino acids that interact with the DNA, was determined and nearly all of them are observed to lose specificity across the entire binding site. The change of Asn at position 460 to His, which corresponds to the natural amino acid at the homologous position in STAT6, does not change the specificity nor does it change the length preference to match that of STAT6. These results provide the first quantitative analysis of changes in binding affinity for the STAT1 protein, and several variants of it, to hundreds of different binding sites including different spacer lengths, and the effect of CpG methylation.
Assuntos
Ilhas de CpG/genética , DNA/genética , Variação Genética , Fator de Transcrição STAT1/genética , Algoritmos , Sequência de Aminoácidos , Sequência de Bases , Sítios de Ligação/genética , Ligação Competitiva , DNA/metabolismo , Metilação de DNA , Eletroforese em Gel de Poliacrilamida , Cinética , Mutação de Sentido Incorreto , Ligação Proteica , Fator de Transcrição STAT1/metabolismo , Fator de Transcrição STAT6/genética , Fator de Transcrição STAT6/metabolismo , Homologia de Sequência de AminoácidosRESUMO
Several variants of a nucleic acid binding motif (RRM1) of putative transcription factor hnRNP LL containing nucleobase amino acids at specific positions have been prepared and used to study binding affinity for the BCL2 i-motif DNA. Molecular modeling suggested a number of amino acids in RRM1 likely to be involved in interaction with the i-motif DNA, and His24 and Arg26 were chosen for modification based on their potential ability to interact with G14 of the i-motif DNA. Four nucleobase amino acids were introduced into RRM1 at one or both of positions 24 and 26. The introduction of cytosine nucleobase 2 into position 24 of RRM1 increased the affinity of the modified protein for the i-motif DNA, consistent with the possible Watson-Crick interaction of 2 and G14. In comparison, the introduction of uracil nucleobase 3 had a minimal effect on DNA affinity. Two structurally simplified nucleobase analogues (1 and 4) lacking both the N-1 and the 2-oxo substituents were also introduced in lieu of His24. Again, the RRM1 analogue containing 1 exhibited enhanced affinity for the i-motif DNA, while the protein analogue containing 4 bound less tightly to the DNA substrate. Finally, the modified protein containing 1 in lieu of Arg26 also bound to the i-motif DNA more strongly than the wild-type protein, but a protein containing 1 both at positions 24 and 26 bound to the DNA less strongly than wild type. The results support the idea of using nucleobase amino acids as protein constituents for controlling and enhancing DNA-protein interaction. Finally, modification of the i-motif DNA at G14 diminished RRM1-DNA interaction, as well as the ability of nucleobase amino acid 1 to stabilize RRM1-DNA interaction.
Assuntos
Aminoácidos/química , DNA/química , Proteínas Proto-Oncogênicas c-bcl-2/química , Sítios de Ligação , Humanos , Modelos Moleculares , Estrutura Molecular , Motivos de NucleotídeosRESUMO
Incorporating information about DNA structure can increase the reliability of predictions of transcription factor binding sites.
Assuntos
DNA/química , Sítios de Ligação , Ligação Proteica , Reprodutibilidade dos Testes , Fatores de TranscriçãoRESUMO
The recently discovered role of the BCL2 (B-cell lymphoma 2 gene) promoter i-motif DNA in modulation of gene expression via interaction with the ribonucleoprotein hnRNP L-like (hnRNP LL) has prompted a more detailed study of the nature of this protein-DNA interaction. The RNA recognition motifs (RRMs) of hnRNP LL were expressed individually, and both RRM1 and RRM2 were found to bind efficiently to the BCL2 i-motif DNA, as well as being critical for transcriptional activation, whereas RRM3-4 bound only weakly to this DNA. Binding was followed by unfolding of the DNA as monitored by changes in the CD spectrum. Mutational analysis of the i-motif DNA revealed that binding involved primarily the lateral loops of the i-motif. The kinetics of binding of the DNA with RRM1 was explored by recording CD spectra at predetermined times following admixture of the protein and DNA. The change in molar ellipticity was readily apparent after 30 s and largely complete within 1 min. A more detailed view of protein-DNA interaction was obtained by introducing the fluorescence donor 6-CNTrp in RRM1 at position 137, and the acceptor 4-aminobenzo[g]quinazoline-2-one (Cf) in lieu of cytidine22 in the i-motif DNA. The course of binding of the two species was monitored by FRET, which reflected a steady increase in energy transfer over a period of several minutes. The FRET signal could be diminished by the further addition of (unlabeled) RRM2, no doubt reflecting competition for binding to the i-motif DNA. These experiments using the individual RRM domains from hnRNP LL confirm the role of this transcription factor in activation of BCL2 transcription via the i-motif in the promoter element.
Assuntos
DNA/genética , DNA/metabolismo , Ribonucleoproteínas Nucleares Heterogêneas/química , Ribonucleoproteínas Nucleares Heterogêneas/metabolismo , Motivos de Nucleotídeos , Regiões Promotoras Genéticas/genética , Proteínas Proto-Oncogênicas c-bcl-2/genética , Sequência de Bases , DNA/química , Modelos Moleculares , Ligação Proteica , Domínios ProteicosRESUMO
Described herein are the syntheses and photophysical characterization of three novel cyanotryptophans, and their efficient incorporation into proteins as fluorescent probes. Photophysical characteristics indicated that each was significantly brighter and red-shifted in fluorescence emission relative to tryptophan. Each analogue was used to activate a suppressor tRNA transcript and was incorporated with good efficiency into two different positions (Trp22 and Trp74) of Escherichia coli dihydrofolate reductase (ecDHFR). The Trp analogues could be monitored selectively in the presence of multiple native Trp residues in DHFR. 6-CNTrp (A) formed an efficient Förster resonance energy transfer (FRET) pair with l-(7-hydroxycoumarin-4-yl)ethylglycine (HCO, D) at position 17. Further, 6-CNTrp (A) was incorporated into two DNA binding proteins, including the Klenow fragment of DNA polymerase I and an RNA recognition motif (RRM2) of heterogeneous nuclear ribonucleoprotein L-like (hnRNP LL). Using these proteins, we demonstrated the use of FRET involving A as a fluorescence donor and benzo[g]quinazoline-2,4-(1H,3H)-dione 2'-deoxyriboside (Tf) or 4-aminobenzo[g]quinazoline-2-one 2'-deoxyriboside (Cf) as fluorescent acceptors to study the binding interaction of the Klenow fragment with duplex DNA oligomers (labeled with Tf), or the domain-specific association between hnRNP LL and the BCL2 i-motif DNA (labeled with Cf). Thus, the non-natural amino acid could be used as a FRET partner for studying protein-nucleic acid interactions. Together, these findings demonstrate the potential utility of 6-CNTrp (A) as a fluorescence donor for the study of protein conformational events.
Assuntos
DNA/química , Corantes Fluorescentes/química , Proteínas/química , Triptofano/análogos & derivados , Transferência Ressonante de Energia de Fluorescência , Ligação Proteica , Conformação Proteica , Triptofano/químicaRESUMO
In an earlier study, ß³-puromycin was used for the selection of modified ribosomes, which were utilized for the incorporation of five different ß-amino acids into Escherichia coli dihydrofolate reductase (DHFR). The selected ribosomes were able to incorporate structurally disparate ß-amino acids into DHFR, in spite of the use of a single puromycin for the selection of the individual clones. In this study, we examine the extent to which the structure of the ß³-puromycin employed for ribosome selection influences the regio- and stereochemical preferences of the modified ribosomes during protein synthesis; the mechanistic probe was a single suppressor tRNA(CUA) activated with each of four methyl-ß-alanine isomers (1-4). The modified ribosomes were found to incorporate each of the four isomeric methyl-ß-alanines into DHFR but exhibited a preference for incorporation of 3(S)-methyl-ß-alanine (ß-mAla; 4), i.e., the isomer having the same regio- and stereochemistry as the O-methylated ß-tyrosine moiety of ß³-puromycin. Also conducted were a selection of clones that are responsive to ß²-puromycin and a demonstration of reversal of the regio- and stereochemical preferences of these clones during protein synthesis. These results were incorporated into a structural model of the modified regions of 23S rRNA, which included in silico prediction of a H-bonding network. Finally, it was demonstrated that incorporation of 3(S)-methyl-ß-alanine (ß-mAla; 4) into a short α-helical region of the nucleic acid binding domain of hnRNP LL significantly stabilized the helix without affecting its DNA binding properties.
Assuntos
Alanina/análogos & derivados , Proteínas de Escherichia coli/biossíntese , Ribonucleoproteínas Nucleares Heterogêneas Grupo L/biossíntese , Modelos Moleculares , RNA Bacteriano/metabolismo , RNA Ribossômico/metabolismo , Ribossomos/metabolismo , Tetra-Hidrofolato Desidrogenase/biossíntese , Alanina/química , Alanina/metabolismo , Escherichia coli/enzimologia , Escherichia coli/metabolismo , Proteínas de Escherichia coli/química , Ribonucleoproteínas Nucleares Heterogêneas Grupo L/química , Ribonucleoproteínas Nucleares Heterogêneas Grupo L/genética , Humanos , Ligação de Hidrogênio , Simulação de Dinâmica Molecular , Proteínas Mutantes/biossíntese , Proteínas Mutantes/química , Proteínas Mutantes/genética , Motivos de Nucleotídeos , Peptidil Transferases/genética , Peptidil Transferases/metabolismo , Conformação Proteica , Estabilidade Proteica , Puromicina/análogos & derivados , Puromicina/química , Puromicina/metabolismo , RNA Bacteriano/química , RNA Ribossômico/química , RNA Ribossômico 23S/química , RNA Ribossômico 23S/metabolismo , Aminoacil-RNA de Transferência/química , Aminoacil-RNA de Transferência/metabolismo , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Estereoisomerismo , Especificidade por Substrato , Tetra-Hidrofolato Desidrogenase/químicaRESUMO
The donor-acceptor (D-A) type dipolar fluorophores, an important class of luminescent dyes with two-photon absorption behaviour, generally emit strongly in organic solvents but poorly in aqueous media. To understand and enhance the poor emission behaviour of dipolar dyes in aqueous media, we undertake a rational approach that includes a systematic structure variation of the donor, amino substituent of acedan, an important two-photon dye. We identify several factors that influence the emission behaviour of the dipolar dyes in aqueous media through computational and photophysical studies on new acedan derivatives. As a result, we can make acedan dyes emit bright fluorescence under one- and two-photon excitation in aqueous media by suppressing the liable factors for poor emission: 1,3-allylic strain, rotational freedom, and hydrogen bonding with water. We also validate that these findings can be generally extended to other dipolar fluorophores, as demonstrated for naphthalimide, coumarin and (4-nitro-2,1,3-benzoxadiazol-7-yl)amine (NBD) dyes. The new acedan and naphthalimide dyes thus allow us to obtain much brighter two-photon fluorescent images in cells and tissues than in their conventional forms. As an application of these findings, a thiol probe is synthesized based on a new naphthalimide dye, which shows greatly enhanced fluorescence from the widely used N,N-dimethyl analogue. The results disclosed here provide essential guidelines for the development of efficient dipolar dyes and fluorescence probes for studying biological systems, particularly by two-photon microscopy.
RESUMO
In a recent study, we described the enhanced double-strand cleavage of hairpin DNAs by Fe·bleomycin (Fe·BLM) that accompanies increasingly strong binding of this antitumor agent and suggested that this effect may be relevant to the mechanism by which BLM mediates its antitumor effects. Because the DNA in tumor cells is known to be hypomethylated on cytidine relative to that in normal cells, it seemed of interest to study the possible effects of methylation status on BLM-induced double-strand DNA cleavage. Three hairpin DNAs found to bind strongly to bleomycin, and their methylated counterparts, were used to study the effect of methylation on bleomycin-induced DNA degradation. Under conditions of limited DNA cleavage, there was a significant overall decrease in the cleavage of methylated hairpin DNAs. Cytidine methylation was found to result in decreased BLM-induced cleavage at the site of methylation and to result in enhanced cleavage at adjacent nonmethylated sites. For two of the three hairpin DNAs studied, methylation was accompanied by a dramatic decrease in the binding affinity for Fe·BLM, suggesting the likelihood of diminished double-strand cleavage. The source of the persistent binding of BLM by the third hairpin DNA was identified. Also identified was the probable molecular mechanism for diminished binding and cleavage of the methylated DNAs by BLM. The possible implications of these findings for the antitumor selectivity of bleomycin are discussed.
Assuntos
Antineoplásicos/farmacologia , Bleomicina/farmacologia , Metilação de DNA , DNA/química , Conformação de Ácido Nucleico , Antineoplásicos/química , Sítios de Ligação , Bleomicina/química , DNA/metabolismo , Sequências Repetidas Invertidas , Modelos Moleculares , Estrutura MolecularRESUMO
Bleomycins A5 and B2 were used to study the structural features in hairpin DNAs conducive to strong BLM-DNA interaction. Two members of a 10-hairpin DNA library previously found to bind most tightly to these BLMs were subsequently noted to share the sequence 5'-ACGC (complementary strand sequence 5'-GCGT). Each underwent double-strand cleavage at five sites within, or near, an eight base pair region of the DNA duplex which had been randomized to create the original library. A new hairpin DNA library was selected based on affinity for immobilized Fe(III)·BLM A5. Two of the 30 newly identified DNAs also contained the sequence 5'-ACGC/5'-GCGT. These DNAs bound to the Fe(II)·BLMs more tightly than any DNA characterized previously. Surface plasmon resonance confirmed tight Fe(III)·BLM B2 binding and gave an excellent fit for a 1:1 binding model, implying the absence of significant secondary binding sites. Fe(II)·BLM A5 was used to assess sites of double-strand DNA cleavage. Both hairpin DNAs underwent double-strand cleavage at five sites within or near the original randomized eight base region. For DNA 12, four of the five double-strand cleavages involved independent single-strand cleavage reactions; DNA 13 underwent double-strand DNA cleavage by independent single-strand cleavages at all five sites. DNA 14, which bound Fe·BLM poorly, was converted to a strong binder (DNA 15) by insertion of the sequence 5'-ACGC/5'-GCGT. These findings reinforce the idea that tighter DNA binding by Fe·BLM leads to increased double-strand cleavage by a novel mechanism and identify a specific DNA motif conducive to strong BLM binding and cleavage.
Assuntos
Bleomicina/química , DNA/química , Compostos Ferrosos/química , Sequência de Bases , Sítios de Ligação , Bleomicina/farmacologia , DNA/efeitos dos fármacos , Clivagem do DNA/efeitos dos fármacos , Compostos Ferrosos/farmacologia , Conformação MolecularRESUMO
As part of our ongoing efforts to identify compounds having potential utility in treating neurodegenerative and mitochondrial disorders, a series of pyridinol analogues have been prepared. The synthetic route employed for the preparation of the new analogues is different, and considerably more efficient, than that used in previously reported studies. The new route yields a pair of pyridinol regioisomers that can be readily separated and evaluated. Their ability to quench lipid peroxidation and reactive oxygen species (ROS), and to preserve mitochondrial membrane potential (Δψm) and support ATP synthesis is reported. The optimal side chain length was found to be 16 carbon atoms. The metabolic stability of those compounds having optimal biological activities was evaluated in vitro using bovine liver microsomes. The omission of any side chain hydroxyl group and introduction of an azetidine moiety at position 6 of the pyridinol redox core (8 and 9) increased their microsomal stability as compared to the exocyclic dimethylamino group. The favorable metabolic stability conferred by the azetidine moiety in compounds 8 and 9 makes these compounds excellent candidates for further evaluation.
Assuntos
Antioxidantes/farmacologia , Citoproteção/efeitos dos fármacos , Doenças Mitocondriais/tratamento farmacológico , Doenças Neurodegenerativas/tratamento farmacológico , Fármacos Neuroprotetores/farmacologia , Piridinas/farmacologia , Animais , Antioxidantes/síntese química , Antioxidantes/química , Bovinos , Células Cultivadas , Relação Dose-Resposta a Droga , Humanos , Peroxidação de Lipídeos/efeitos dos fármacos , Linfócitos/efeitos dos fármacos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Microssomos Hepáticos/química , Microssomos Hepáticos/metabolismo , Doenças Mitocondriais/patologia , Estrutura Molecular , Doenças Neurodegenerativas/patologia , Fármacos Neuroprotetores/síntese química , Fármacos Neuroprotetores/química , Piridinas/síntese química , Piridinas/química , Espécies Reativas de Oxigênio/antagonistas & inibidores , Espécies Reativas de Oxigênio/metabolismo , Relação Estrutura-AtividadeRESUMO
Since the early nineties there have been a number of reports on the experimental development of Mg electrolytes based on organo/amide-magnesium chlorides and their transmetalations. However, there are no theoretical papers describing the underlying operating mechanisms of Mg electrolytes, and there is no clear understanding of these mechanisms. We have therefore attempted to clarify the operating mechanisms of Mg electrolytes by studying the characteristics of Mg complexes, solvation, chemical equilibrium, Mg-deposition processes, electrolyte-oxidation processes, and oxidative degradation mechanism of RMgCl-based electrolytes, using ab initio calculations. The formation and solvation energies of Mg complexes highly depend on the characteristics of R groups. Thus, changes in R groups of RMgCl lead to changes in the equilibrium position and the electrochemical reduction and oxidation pathways and energies. We first provide a methodological scheme for calculating Mg reduction potential values in non-aqueous electrolytes and electrochemical windows. We also describe a strategy for designing Mg electrolytes to maximize the electrochemical windows and oxidative stabilities. These results will be useful not only for designing improved Mg electrolytes, but also for developing new electrolytes in the future.
RESUMO
Clinically used bleomycin A5 has been employed in a study of double-strand cleavage of a library of 10 hairpin DNAs originally selected on the basis of their strong binding to bleomycin. Each of the DNAs underwent double-strand cleavage at more than one site, and all of the cleavage sites were within, or in close proximity to, an eight-base-pair region of the duplex that had been randomized to create the original library. A total of 31 double-strand cleavage sites were identified on the 10 DNAs, and 14 of these sites were found to represent coupled cleavage sites, that is, events in which one of the two strands was always cleaved first, followed by the associated site on the opposite strand. Most of these coupled sites underwent cleavage by a mechanism described previously by the Povirk laboratory and afforded cleavage patterns entirely analogous to those reported. However, at least one coupled cleavage event was noted that did not conform to the pattern of those described previously. More surprisingly, 17 double-strand cleavages were found not to result from coupled double-strand cleavage, and we posit that these cleavages resulted from a new mechanism not previously described. Enhanced double-strand cleavages at these sites appear to be a consequence of the dynamic nature of the interaction of Fe·BLM A5 with the strongly bound hairpin DNAs.
Assuntos
Bleomicina/química , DNA/química , Sequência de Bases , Quebras de DNA de Cadeia Dupla , Clivagem do DNA , Conformação MolecularRESUMO
A study of BLM A5 was conducted using a previously isolated library of hairpin DNAs found to bind strongly to metal-free BLM. The ability of Fe(II)·BLM to affect cleavage on both the 3' and 5' arms of the hairpin DNAs was characterized. The strongly bound DNAs were found to be efficient substrates for Fe·BLM A5-mediated hairpin DNA cleavage. Surprisingly, the most prevalent site of BLM-mediated cleavage was found to be the 5'-AT-3' dinucleotide sequence. This dinucleotide sequence and other sequences generally not cleaved well by BLM when examined using arbitrarily chosen DNA substrates were apparent when examining the library of 10 hairpin DNAs. In total, 132 sites of DNA cleavage were produced by exposure of the hairpin DNA library to Fe·BLM A5. The existence of multiple sites of cleavage on both the 3' and 5' arms of the hairpin DNAs suggested that some of these might be double-strand cleavage events. Accordingly, an assay was developed to test the propensity of the hairpin DNAs to undergo double-strand DNA damage. One hairpin DNA was characterized using this method and gave results consistent with earlier reports of double-strand DNA cleavage but with a sequence selectivity that was different from those reported previously.
Assuntos
Antibióticos Antineoplásicos/farmacologia , Bleomicina/farmacologia , DNA/química , Biblioteca Gênica , Antibióticos Antineoplásicos/química , Sequência de Bases , Bleomicina/química , DNA/genética , Clivagem do DNA/efeitos dos fármacos , Sequências Repetidas Invertidas/efeitos dos fármacos , Dados de Sequência MolecularRESUMO
A series of aza analogues (4-9) of the experimental neuroprotective drug idebenone (1) have been prepared and evaluated for their ability to attenuate oxidative stress induced by glutathione depletion and to compensate for the decrease in oxidative phosphorylation efficiency in cultured Friedreich's ataxia (FRDA) fibroblasts and lymphocytes and also coenzyme Q10-deficient lymphocytes. Modification of the redox core of the previously reported 3 improved its antioxidant and cytoprotective properties. Compounds 4-9, having the same redox core, exhibited a range of antioxidant activities, reflecting side chain differences. Compounds having side chains extending 14-16 atoms from the pyrimidinol ring (6, 7, and 9) were potent antioxidants. They were superior to idebenone and more active than 3, 4, 5, and 8. Optimized analogue 7 and its acetate (7a) are of interest in defining potential therapeutic agents capable of blocking oxidative stress, maintaining mitochondrial membrane integrity, and augmenting ATP levels. Compounds with such properties may find utility in treating mitochondrial and neurodegenerative diseases such as FRDA and Alzheimer's disease.
RESUMO
Mononuclear Zn(II)-DPA and Cu(II)-DPA complexes crafted on 2-hydroxy-6-cyanonaphthalene fluorophore selectively recognize PPi over ATP and other anions including inorganic phosphates in aqueous medium, showing turn-on type fluorescence enhancements. Coordination of a hydroxyl group of the fluorophore, directly or in alkoxy form, to the central metal ion is crucial for the sensing processes. Both the complexes elicit a fluorescence increase in a time-dependent fashion.
Assuntos
Aminas/química , Difosfatos/análise , Corantes Fluorescentes/síntese química , Sondas Moleculares/síntese química , Naftóis/química , Compostos Organometálicos/síntese química , Ácidos Picolínicos/química , Ânions , Cobre/química , Cobre/metabolismo , Fluorescência , Corantes Fluorescentes/metabolismo , Sondas Moleculares/metabolismo , Compostos Organometálicos/metabolismo , Fosfatos/química , Soluções , Espectrometria de Fluorescência , Água , Zinco/química , Zinco/metabolismoRESUMO
A ratiometric fluorescent probe for mercury species is developed based on the metal-promoted hydrolysis of a vinyl ether derivative of 2-(benzothiazol-2-yl)phenol in a buffer solution. The probe responds selectively to mercury species over various other metal ions with a marked fluorescence change from blue to cyan through the excited state intramolecular proton transfer (ESIPT) process. The fluorescence titration is complete with 0.5 equiv of HgCl(2), which indicates that the probe also responds to organomercury species, RHgCl.
Assuntos
Acetanilidas/química , Benzotiazóis/química , Corantes Fluorescentes/química , Mercúrio/química , Hidrólise , Estrutura MolecularRESUMO
Chemical probes are valuable tools for the investigation of biochemical processes, diagnosis of disease markers, detection of hazardous compounds, and other purposes. Therefore, the development of chemical probes continues to grow through various approaches with different disciplines and design strategies. Fluorescent probes have received much attention because they are sensitive and easy-to-operate, in general. To realize desired selectivity toward a given analyte, the recognition site of a fluorescent probe is designed in such a way to maximize the binding interactions, usually through weak molecular forces such as hydrogen bonding, toward the analyte over other competing ones. In addition to such a supramolecular approach, the development of fluorescent probes that sense analytes through chemical reactions has witnessed its usefulness for achieving high selectivity, in many cases, superior to that obtainable by the supramolecular approach. Creative incorporations of the reactive groups to latent fluorophores have provided novel chemical probes for various analytes. In this feature article, we overview the recent progress in the development of turn-on fluorescent probes that are operating through chemical reactions triggered by target analytes. Various chemical reactions have been implemented in the development of many reactive probes with very high selectivity and sensitivity toward target analytes. A major emphasis has been focused on the type of chemical reactions utilized, with the hope that further explorations can be made with new chemical reactions to develop reactive probes useful for various applications.
Assuntos
Corantes Fluorescentes/química , Catálise , Metais/química , Espectrometria de Fluorescência/métodosRESUMO
A new class of chimeric molecules have been developed. These are based on polyphenols like catechin and epicatechin and monocyclic beta-lactams. The two units are joined via a triazole linker using the 'Click Chemistry' conditions. The compounds showed good to weak antibacterial activity against Escherichia coli as well as moderate inhibition of RNase A.