The mechanism of antimalarial action of the ruthenium-chloroquine complex [RuCl 2 (CQ)] 2 (1), previously shown by us to be active in vitro against CQ-resistant strains of Plasmodium falciparum and in vivo against P. berghei, has been investigated. The complex is rapidly hydrolyzed in aqueous solution to [RuCl(OH 2 ) 3 (CQ)] 2 [Cl] 2 , which is probably the active species. This compound binds to hematin in solution and inhibits aggregation to β-hematin at pH ∼ 5 to a slightly lower extent than chloroquine diphosphate; more importantly, the heme aggregation inhibition activity of complex 1 is significantly higher than that of CQ when measured at the interface of noctanol-aqueous acetate buffer mixtures under acidic conditions modeling the food vacuole of the parasite. Partition coefficient measurements confirmed that complex 1 is considerably more lipophilic than CQ in n-octanol-water mixtures at pH ∼ 5. This suggests that the principal target of complex 1 is the heme aggregation process, which has recently been reported to be fast and spontaneous at or near water-lipid interfaces. The enhanced antimalarial activity of complex 1 is thus probably due to a higher effective concentration of the drug at or near the interface compared with that of CQ, which accumulates strongly in the aqueous regions of the vacuole under those conditions. Furthermore, the activity of complex 1 against CQ-resistant strains of P. falciparum is probably related to its greater lipophilicity, in line with previous reports indicating a lowered ability of the mutated transmembrane transporter PfCRT to promote the efflux of highly lipophilic drugs. The metal complex also interacts with DNA by intercalation, to a comparable extent and in a similar manner to uncomplexed CQ and therefore DNA binding does not appear to be an important part of the mechanism of antimalarial action in this case.

We have constructed a fiber optic device that internally flows triplet oxygen and externally produces singlet oxygen, causing a reaction at the (Z)-1,2-dialkoxyethene spacer group, freeing a pheophorbide sensitizer upon the fragmentation of a reactive dioxetane intermediate. The device can be operated and sensitizer photorelease observed using absorption and fluorescence spectroscopy. We demonstrate the preference of sensitizer photorelease when the probe tip is in contact with octanol or lipophilic media. A first-order photocleavage rate constant of 1.13 h−1 was measured in octanol where dye desorption was not accompanied by readsorption. When the probe tip contacts aqueous solution, the photorelease was inefficient because most of the dye adsorbed on the probe tip, even after the covalent ethene spacer bonds have been broken. The observed stability of the free sensitizer in lipophilic media is reasonable even though it is a pyropheophorbide-a derivative that carries a p-formylbenzylic alcohol substituent at the carboxylic acid group. In octanol or lipid systems, we found that the dye was not susceptible to hydrolysis to pyropheophorbide-a, otherwise a pH effect was observed in a binary methanol-water system (9:1) at pH below 2 or above 8.

The solvatochromism and possible excited state deactivation pathways of benzo[a]fluorenone were investigated and compared with the related aromatic compounds 9‐fluorenone and benzo[b]fluorenone. These molecules can serve as molecular building blocks with application in light emitting diodes, catalysts, and dye‐sensitized solar cells. A study of the effect changes in aromatic structure might have on their relative photophysical responses is therefore of interest. Steady‐state absorption and emission spectra, Lippert‐Mataga plots, fluorescence lifetimes, and absolute quantum yields were used in tandem with time dependent density functional theory (TD‐DFT) calculations to investigate solvatochromism, excited state decay, and the relative energies of singlet and triplet excited states. It was found that changes in aromaticity affect the energetic order and character of the excited singlet and triplet states differently among these fluorenones as solvent polarity is varied. The observed fluorescence emission from all these molecules can be understood in terms of their relative ability to undergo intersystem crossing. Additional calculations on the related benzo[c]fluorenone predict that its solvatochromic behavior should resemble that of benzo[a]fluorenone.

No abstract

Short double-stranded RNA molecules have recently emerged as important regulators of gene expression. These small RNAs associate with a member of the Argonaute protein family in an assembly known as RNA-induced silencing complex (RISC).Here we elucidate the pathway of RNA Interference (RNAi) in vivo by applying fluorescence correlation and cross-correlation spectroscopy (FCS/FCCS). We show that two distinct RISC exist: a large ~3 MDa complex in the cytoplasm and a 20-fold smaller complex in the nucleus. Nuclear RISC, consisting only of Ago2 and a short RNA, is loaded in the cytoplasm and imported into the nucleus. The import of Ago2 into the nucleus is mediated by the import receptor Impotin8. We further demonstrate that FCCS can be used to study the interaction of different members of the Argonaute protein family with short double-stranded RNAs and their target mRNA molecules.

DNA G-quadruplexes are enriched near the transcription start site (TSS) of the human genes and have been suggested to be involved in gene transcription and translation. Informations about G-quadruplex conformation and dynamics is crucial to our understanding of the roles of quadruplex in gene regulation as well as to the development of novel therapeutic agents that interact with the quadruplex therefore modulate gene expression. Single molecule fluorescence resonance energy transfer (smFRET) can resolve conformational heterogeneity and dynamic fluctuations in nucleic acids, providing unique insights into the biophysics of quadruplex. We have recently elucidated the conformational heterogeneity and dynamics of the quadruplexes formed in the promoter of human c-myc and c-kit genes by smFRET. Here we present single molecule analysis of DNA quadruplex elements found in the TSS of the promoter of the MEF2D, a member of MEF2 (myocyte enhancer factor-2) family of transcription factors which regulate the response of heart to cardiac stress signals, and also in the chromosome 19 specific minisatellite sequences in the promoter of human cardiac troponin I (TNNI3), a gene encodes constituent protein of the troponin complex on the thin filament of cardiac muscle.

revealed observable and quantifiable conformational changes at the ATP and RNA binding sites. Trajectories of minimum distances among residues, and model-to-model ratios of RMSF were used as parameters to quantify the structural effects of the mutations. Atomic interactions propagated from the mutational residues to both ATP and RNA binding sites were observed due to the changes of side chains of these mutations. We hypothesize these three residues play structural roles in the unique functions of DbpA in rRNA interactions. Future studies targeting key unique residues may reveal connected allosteric pathways within the protein structure and assist studying the diversity of function among DEAD-box proteins.

No abstract