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Resonance Energy Transfer (resonance + energy_transfer)
Kinds of Resonance Energy Transfer Selected AbstractsNanoparticle Electroluminescence: Controlling Emission Color Through Förster Resonance Energy Transfer in Hybrid ParticlesADVANCED FUNCTIONAL MATERIALS, Issue 22 2009Christopher F. Huebner Abstract Electroluminescent (EL) polymers are attractive for developing all-organic light-emitting devices (OLEDs) due to the potential advantages that polymeric systems may offer in the large-scale manufacturing of electronics. Nonetheless, many of these EL , -conjugated polymers are inherently insoluble in the solvents employed in the intended solution-based manufacturing processes. One such polymer is poly(2,5-dioctyl-1,4-phenylenevinylene) (POPPV), where the inherent lack of solubility of POPPV in organic solvents has frustrated its widespread application in devices and no OLEDs have been presented that exploit its electroluminescence characteristics. In this effort, a unique strategy is presented for the preparation of hybrid nanoparticles composed of POPPV, a green emitter (,em,=,505,nm) and poly(9,9-di- n -octylfluorenyl-2,7-diyl) (PFO), a blue emitter (,em,=,417,nm). The aqueous-based nanoparticle dispersion composed of these hybrid particles is stable to aggregate and can be employed in the construction of OLEDs. The color characteristics of the electroluminescence for the devices can be tuned by exploiting the Förster resonance energy transfer between the polymers within a particle, while suppressing energy transfer between the particles. These aqueous-based nanoparticle dispersions are amenable to being printed into devices through high-throughput manufacturing techniques, for example, roll-to-roll printing. [source] Two-Photon Excitation of Quantum-Dot-Based Fluorescence Resonance Energy Transfer and Its Applications,ADVANCED MATERIALS, Issue 15 2007R. Clapp Nonradiative fluorescence resonance energy transfer (FRET) between a luminescent quantum dot (QD) donor and a proximal dye brought in close proximity of the QD surface via conjugation with a dye-labeled peptide (or a protein) is shown (see figure). The system is excited with near IR irradiation (well below the absorption band of the QD), via a fast two-photon process, which produces a FRET signal with very low background contribution due to a substantially reduced nonlinear direct excitation of the dye. [source] The ,670A > G polymorphism in the promoter region of the FAS gene is associated with necrosis in periportal areas in patients with chronic hepatitis CJOURNAL OF VIRAL HEPATITIS, Issue 6 2005J. Aguilar-Reina Summary., Evidence suggests that apoptosis of liver cells may play a significant role in the pathogenesis of hepatitis C virus (HCV) infection. One of the best characterized apoptotic pathway is that mediated by the death receptor Fas. Fas expression has been found to be up-regulated on hepatocytes in chronic HCV infection, particularly in periportal areas. Recently, two polymorphisms have been identified in the promotor region of the FAS gene, ,1377G > A and ,670A > G. We have evaluated the involvement of these variants in the susceptibility to HCV infection, the severity of liver damage and progression of fibrosis in chronic hepatitis C. A cohort of 197 patients with chronic hepatitis C and 100 controls were analysed for both polymorphisms by Fluorescence Resonance Energy Transfer using specific probes and the LightcyclerTM system. In addition, liver biopsies were taken in 167 patients and scored using the Knodell classification system. We have found that the allele frequencies and the distribution of both polymorphisms do not differ significantly in the HCV cohort and in the control population. Thus, none of the polymorphisms seems to be related with susceptibility to HCV infection. However, we have examined the possible association between the two variants and the grade of necroinflammatory activity and the stage of fibrosis and we have detected an under-representation of the ,670A > G variant among those patients with higher Knodell's scores (P = 0.049) and necroinflammatory activity (P = 0.036). The ,670A allele was associated with higher levels of periportal necrosis (P = 0.012). In conclusion, our findings suggest an association between the ,670A > G polymorphism and the grade of necrosis in periportal areas in patients with chronic hepatitis C. [source] Detection of protease activity by FRET using porous silicon as an energy acceptorPHYSICA STATUS SOLIDI (A) APPLICATIONS AND MATERIALS SCIENCE, Issue 6 2009Luo Gu Abstract This work demonstrates a method for detection of protease activity using porous Si as the energy acceptor in a Fluorescence Resonance Energy Transfer (FRET) assay. A fluorescent dye (fluorescein) is trapped in a porous Si matrix, where its fluorescence is quenched. The dye is trapped using a protein (zein), and the action of proteases (Pronase E) release the dye from the quenching matrix. A strong fluorescence signal appears within 30 min of Pronase E addition. A control with no Pronase E shows little fluorescence, and a control using heat-denatured Pronase E shows approximately 50% of the fluorescence of the active protease. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) [source] Pentiptycene-Derived Oligo(p -phenyleneethynylene)s: Conformational Control, Chain-Length Effects, Localization of Excitation, and Intrachain Resonance Energy Transfer,ANGEWANDTE CHEMIE, Issue 52 2009Jye-Shane Yang Prof. Ein lokalisierter angeregter Zustand resultiert, weil Pentiptycen-Pentiptycen-Diederwinkel in von Pentiptycen abgeleiteten Oligo(p -phenylenethinylenen) bevorzugt groß sind und die Relaxation der Verdrillung in 2-Methyltetrahydrofuran-Glas bei 80,K eingeschränkt ist. Damit ist nicht nur das Konzept der torsionsinduzierten Lokalisierung einer Anregung, sondern auch die effiziente Intraketten-Energieübertragung in konjugierten Polymeren demonstriert. [source] Marked changes in signal transduction upon heteromerization of dopamine D1 and histamine H3 receptorsBRITISH JOURNAL OF PHARMACOLOGY, Issue 1 2009Carla Ferrada Background and purpose:, Functional interactions between the G protein-coupled dopamine D1 and histamine H3 receptors have been described in the brain. In the present study we investigated the existence of D1,H3 receptor heteromers and their biochemical characteristics. Experimental approach:, D1,H3 receptor heteromerization was studied in mammalian transfected cells with Bioluminescence Resonance Energy Transfer and binding assays. Furthermore, signalling through mitogen-activated protein kinase (MAPK) and adenylyl cyclase pathways was studied in co-transfected cells and compared with cells transfected with either D1 or H3 receptors. Key results:, Bioluminescence Resonance Energy Transfer and binding assays confirmed that D1 and H3 receptors can heteromerize. Activation of histamine H3 receptors did not lead to signalling towards the MAPK pathway unless dopamine D1 receptors were co-expressed. Also, dopamine D1 receptors, usually coupled to Gs proteins and leading to increases in cAMP, did not couple to Gs but to Gi in co-transfected cells. Furthermore, signalling via each receptor was blocked not only by a selective antagonist but also by an antagonist of the partner receptor. Conclusions and implications:, D1,H3 receptor heteromers constitute unique devices that can direct dopaminergic and histaminergic signalling towards the MAPK pathway in a Gs -independent and Gi -dependent manner. An antagonist of one of the receptor units in the D1,H3 receptor heteromer can induce conformational changes in the other receptor unit and block specific signals originating in the heteromer. This gives rise to unsuspected therapeutic potentials for G protein-coupled receptor antagonists. [source] A Nonenzymatic Chemiluminescent Reaction Enabling Chemiluminescence Resonance Energy Transfer to Quantum DotsCHEMISTRY - A EUROPEAN JOURNAL, Issue 21 2010Shulin Zhao Glowing transfer: An efficient chemiluminescence resonance energy transfer (CRET) between a luminol donor and a quantum dot acceptor was observed for the first time from a nonenzymatic reaction involving luminol,NaBrO,CdTe quantum dots (see figure). This work represents significant progress from the previously reported observation of CRET between a luminol donor and a horseradish peroxidase conjugated quantum dot acceptor from an enzyme-catalyzed chemiluminescence reaction. [source] An Efficient Fluorescence Resonance Energy Transfer (FRET) between Pyrene and Perylene Assembled in a DNA Duplex and Its Potential for Discriminating Single-Base ChangesCHEMISTRY - A EUROPEAN JOURNAL, Issue 8 2010Hiromu Kashida Dr. Abstract To increase the apparent Stokes' shift of perylene, pyrene (donor) and perylene (acceptor) were assembled in a DNA duplex to achieve the efficient fluorescence resonance energy transfer (FRET) from pyrene to perylene. Multiple donors were introduced in the vicinity of acceptors through D -threoninol and natural base pairs were inserted between the dyes. Accordingly, donors and acceptors could be accumulated inside the DNA without forming an undesired excimer/exciplex. When two pyrene moieties were located in proximity to one perylene with one base pair inserted between them, efficient FRET occurred within the duplex. Thus, strong emission at 460,nm was observed from perylene when excited at 345,nm at which pyrene has its absorption. The apparent Stokes' shift became as large as 115,nm with a high apparent FRET efficiency (,>1). However, the introduction of more than two pyrenes did not enhance the fluorescence intensity of perylene, due to the short Förster radius (R0) of the donor pyrene. Next, this FRET system was used to enlarge the Stokes' shift of the DNA probe, which can discriminate a one-base deletion mutant from wild type with a model system by incorporation of multiple donors into DNA. Two perylene moieties were tethered to the DNA on both sides of the intervening base, and two pyrenes were further inserted in the vicinity of the perylenes as an antenna. Hybridization of this FRET probe with a fully matched DNA allowed monomer emission of perylene when the pyrenes were excited. In contrast, excimer emission was generated by hybridization with a one-base deletion mutant. Thus, the apparent Stokes' shift was enhanced without loss of efficiency in the detection of the deletion mutant. [source] Nucleosome Immobilization Strategies for Single-Pair FRET Microscopy,CHEMPHYSCHEM, Issue 14 2008Wiepke J. A. Koopmans Abstract All genomic transactions in eukaryotes take place in the context of the nucleosome, the basic unit of chromatin, which is responsible for DNA compaction. Overcoming the steric hindrance that nucleosomes present for DNA-processing enzymes requires significant conformational changes. The dynamics of these have been hard to resolve. Single-pair Fluorescence Resonance Energy Transfer (spFRET) microscopy is a powerful technique for observing conformational dynamics of the nucleosome. Nucleosome immobilization allows the extension of observation times to a limit set only by photobleaching, and thus opens the possibility of studying processes occurring on timescales ranging from milliseconds to minutes. It is crucial however, that immobilization itself does not introduce artifacts in the dynamics. Here we report on various nucleosome immobilization strategies, such as single-point attachment to polyethylene glycol (PEG) or surfaces coated with bovine serum albumin (BSA), and confinement in porous agarose or polyacrylamide gels. We compare the immobilization specificity and structural integrity of immobilized nucleosomes. A crosslinked star polyethylene glycol coating performs best with respect to tethering specificity and nucleosome integrity, and enables us to reproduce for the first time bulk nucleosome unwrapping kinetics in single nucleosomes without immobilization artifacts. [source] Conformations within soluble oligomers and insoluble aggregates revealed by resonance energy transferBIOPOLYMERS, Issue 4 2010Jyothi L. Digambaranath Abstract A fluorescently labeled 20-residue polyglutamic acid (polyE) peptide 20 amino acid length polyglutamic acid (E20) was used to study structural changes which occur in E20 as it co-aggregates with other unlabeled polyE peptides. Resonance energy transfer (RET) was performed using an o -aminobenzamide donor at the N-terminus and 3-nitrotyrosine acceptor at the C-terminus of E20. PolyE aggregates were not defined as amyloid, as they were nonfibrillar and did not bind congo red. Circular dichroism measurements indicate that polyE aggregation involves a transition from ,-helical monomers to aggregated ,-sheets. Soluble oligomers are also produced along with aggregates in the reaction, as determined through size exclusion chromatography. Time-resolved and steady-state RET measurements reveal four dominant E20 conformations: (1) a partially collapsed conformation (24 Å donor,acceptor distance) in monomers, (2) an extended conformation in soluble oligomers (>29 Å donor,acceptor distance), (3) a minor partially collapsed conformation (22 Å donor-acceptor distance) in aggregates, and (4) a major highly collapsed conformation (13 Å donor,acceptor distance) in aggregates. These findings demonstrate the use of RET as a means of determining angstrom-level structural details of soluble oligomer and aggregated states of proteins. © 2009 Wiley Periodicals, Inc. Biopolymers 93: 299,317, 2010. This article was originally published online as an accepted preprint. The "Published Online" date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com [source] Neural agrin increases postsynaptic ACh receptor packing by elevating rapsyn protein at the mouse neuromuscular synapseDEVELOPMENTAL NEUROBIOLOGY, Issue 9 2008Jennifer Brockhausen Abstract Fluorescence resonance energy transfer (FRET) experiments at neuromuscular junctions in the mouse tibialis anterior muscle show that postsynaptic acetylcholine receptors (AChRs) become more tightly packed during the first month of postnatal development. Here, we report that the packing of AChRs into postsynaptic aggregates was reduced in 4-week postnatal mice that had reduced amounts of the AChR-associated protein, rapsyn, in the postsynaptic membrane (rapsyn+/, mice). We hypothesize that nerve-derived agrin increases postsynaptic expression and targeting of rapsyn, which then drives the developmental increase in AChR packing. Neural agrin treatment elevated the expression of rapsyn in C2 myotubes by a mechanism that involved slowing of rapsyn protein degradation. Similarly, exposure of synapses in postnatal muscle to exogenous agrin increased rapsyn protein levels and elevated the intensity of anti-rapsyn immunofluorescence, relative to AChR, in the postsynaptic membrane. This increase in the rapsyn-to-AChR immunofluorescence ratio was associated with tighter postsynaptic AChR packing and slowed AChR turnover. Acute blockade of synaptic AChRs with ,-bungarotoxin lowered the rapsyn-to-AChR immunofluorescence ratio, suggesting that AChR signaling also helps regulate the assembly of extra rapsyn in the postsynaptic membrane. The results suggest that at the postnatal neuromuscular synapse agrin signaling elevates the expression and targeting of rapsyn to the postsynaptic membrane, thereby packing more AChRs into stable, functionally-important AChR aggregates. © 2008 Wiley Periodicals, Inc. Develop Neurobiol, 2008 [source] Study of binding stoichiometries of the human immunodeficiency virus type,1 reverse transcriptase by capillary electrophoresis and laser-induced fluorescence polarization using aptamers as probesELECTROPHORESIS, Issue 2 2006Hao Fu Abstract Binding stoichiometries between four DNA aptamers (RT12, RT26, RTlt49, and ODN93) and the reverse transcriptase (RT) of the type,1 human immunodeficiency virus (HIV-1) were studied using affinity CE (ACE) coupled with LIF polarization and fluorescence polarization (FP). The ACE/LIF study showed evidence of two binding stoichiometries between the HIV-1,RT protein and aptamers RT12, RT26, and ODN93, suggesting that these aptamers can bind to both the p66 and p51 subunits of the HIV-1,RT. Only one binding stoichiometry for aptamer RTlt49 was found. The affinity complexes were easily separated from the unbound aptamers; however, the different stoichiometries were not well resolved. A complementary technique, FP, was able to provide additional information about the binding and supporting evidence for the ACE/LIF results. The ACE/LIFP study also revealed that the FP values of the 1:1 complexes of the HIV-1,RT protein with aptamers RT12, RT26, and ODN93 were always much greater than those of the 1:2 complexes. This was initially surprising because the larger molecular size of the 1:2 complexes was expected to result in higher FP values than the corresponding 1:1 complexes. This phenomenon was probably a result of fluorescence resonance energy transfer between the two fluorescent molecules bound to the HIV-1,RT protein. [source] Role of the plasma membrane leaflets in drug uptake and multidrug resistanceFEBS JOURNAL, Issue 5 2010Hagar Katzir The present study aimed to investigate the role played by the leaflets of the plasma membrane in the uptake of drugs into cells and in their extrusion by P-glycoprotein and multidrug resistance-associated protein 1. Drug accumulation was monitored by fluorescence resonance energy transfer from trimethylammonium-diphenyl-hexatriene (TMA-DPH) located at the outer leaflet to a rhodamine analog. Uptake of dye into cells whose mitochondria had been inactivated was displayed as two phases of TMA-DPH fluorescence quenching. The initial phase comprised a rapid drop in fluorescence that was neither affected by cooling the cells on ice, nor by activity of mitochondria or ABC transporters. This phase reflects the association of dye with the outer leaflet of the plasma membrane. The subsequent phase of TMA-DPH fluorescence quenching occurred in drug-sensitive cell lines with a half-life in the range 20,40 s. The second phase of fluorescence quenching was abolished by incubation of the cells on ice and was transiently inhibited in cells with active mitochondria. Thus, the second phase of fluorescence quenching reflects the accumulation of dye in the cytoplasmic leaflet of the plasma membrane, presumably as a result of flip-flop of dye across the plasma membrane and slow diffusion from the inner leaflet into the cells. Whereas activity of P-glycoprotein prevented the second phase of fluorescence quenching, the activity of multidrug resistance-associated protein 1 had no effect on this phase. Thus, P-glycoprotein appears to pump rhodamines from the cytoplasmic leaflet either to the outer leaflet or to the outer medium. [source] A fluorescence energy transfer-based mechanical stress sensor for specific proteins in situFEBS JOURNAL, Issue 12 2008Fanjie Meng To measure mechanical stress in real time, we designed a fluorescence resonance energy transfer (FRET) cassette, denoted stFRET, which could be inserted into structural protein hosts. The probe was composed of a green fluorescence protein pair, Cerulean and Venus, linked with a stable ,-helix. We measured the FRET efficiency of the free cassette protein as a function of the length of the linker, the angles of the fluorophores, temperature and urea denaturation, and protease treatment. The linking helix was stable to 80 °C, unfolded in 8 m urea, and rapidly digested by proteases, but in all cases the fluorophores were unaffected. We modified the ,-helix linker by adding and subtracting residues to vary the angles and distance between the donor and acceptor, and assuming that the cassette was a rigid body, we calculated its geometry. We tested the strain sensitivity of stFRET by linking both ends to a rubber sheet subjected to equibiaxial stretch. FRET decreased proportionally to the substrate strain. The naked cassette expressed well in human embryonic kidney-293 cells and, surprisingly, was concentrated in the nucleus. However, when the cassette was located into host proteins such ,-actinin, nonerythrocyte spectrin and filamin A, the labeled hosts expressed well and distributed normally in cell lines such as 3T3, where they were stressed at the leading edge of migrating cells and relaxed at the trailing edge. When collagen-19 was labeled near its middle with stFRET, it expressed well in Caenorhabditis elegans, distributing similarly to hosts labeled with a terminal green fluorescent protein, and the worms behaved normally. [source] C-terminal truncated cannabinoid receptor 1 coexpressed with G protein trimer in Sf9 cells exists in a precoupled state and shows constitutive activityFEBS JOURNAL, Issue 23 2007Chandramouli Reddy Chillakuri We have investigated the existence of a precoupled form of the distal C-terminal truncated cannabinoid receptor 1 (CB1-417) and heterotrimeric G proteins in a heterologous insect cell expression system. CB1-417 showed higher production levels than the full-length receptor. The production levels obtained in our expression system were double the values reported in the literature. We also observed that at least the distal C-terminus of the receptor was not involved in receptor dimerization, as was predicted in the literature. Using fluorescence resonance energy transfer, we found that CB1-417 and G,i1,1,2 proteins were colocalized in the cells. GTP,S binding assays with the Sf9 cell membranes containing CB1-417 and the G protein trimer showed that the receptor could constitutively activate the G,i1 protein in the absence of agonists. A CB1-specific antagonist (SR 141716A) inhibited this constitutive activity of the truncated receptor. We found that the CB1-417/G,i1,1,2 complex could be solubilized from Sf9 cell membranes and coimmunoprecipitated. In this study, we have proven that the receptor and G proteins can be coexpressed in higher yields using Sf9 cells, and that the protein complex is stable in detergent solution. Thus, our system can be used to produce sufficient quantities of the protein complex to start structural studies. [source] Effect of mutations in the ,5,,7 loop on the structure and properties of human small heat shock protein HSP22 (HspB8, H11)FEBS JOURNAL, Issue 21 2007Alexei S. Kasakov The human genome encodes ten different small heat shock proteins, each of which contains the so-called ,-crystallin domain consisting of 80,100 residues and located in the C-terminal part of the molecule. The ,-crystallin domain consists of six or seven ,-strands connected by different size loops and combined in two ,-sheets. Mutations in the loop connecting the ,5 and ,7 strands and conservative residues of ,7 in ,A-, ,B-crystallin and HSP27 correlate with the development of different congenital diseases. To understand the role of this part of molecule in the structure and function of small heat shock proteins, we mutated two highly conservative residues (K137 and K141) of human HSP22 and investigated the properties of the K137E and K137,141E mutants. These mutations lead to a decrease in intrinsic Trp fluorescence and the double mutation decreased fluorescence resonance energy transfer from Trp to bis-ANS bound to HSP22. Mutations K137E and especially K137,141E lead to an increase in unordered structure in HSP22 and increased susceptibility to trypsinolysis. Both mutations decreased the probability of dissociation of small oligomers of HSP22, and mutation K137E increased the probability of HSP22 crosslinking. The wild-type HSP22 possessed higher chaperone-like activity than their mutants when insulin or rhodanase were used as the model substrates. Because conservative Lys residues located in the ,5,,7 loop and in the ,7 strand appear to play an important role in the structure and properties of HSP22, mutations in this part of the small heat shock protein molecule might have a deleterious effect and often correlate with the development of different congenital diseases. [source] NEMO oligomerization in the dynamic assembly of the I,B kinase core complexFEBS JOURNAL, Issue 10 2007Elisabeth Fontan NF-,B essential modulator (NEMO) plays an essential role in the nuclear factor ,B (NF-,B) pathway as a modulator of the two other subunits of the I,B kinase (IKK) complex, i.e. the protein kinases, IKK, and IKK,. Previous reports all envision the IKK complex to be a static entity. Using glycerol-gradient ultracentrifugation, we observed stimulus-dependent dynamic IKK complex assembly. In wild-type fibroblasts, the kinases and a portion of cellular NEMO associate in a 350-kDa high-molecular-mass complex. In response to constitutive NF-,B stimulation by Tax, we observed NEMO recruitment and oligomerization to a shifted high-molecular-mass complex of 440 kDa which displayed increased IKK activity. This stimulus-dependent oligomerization of NEMO was also observed using fluorescence resonance energy transfer after a transient pulse with interleukin-1,. In addition, fully activated, dimeric kinases not bound to NEMO were detected in these Tax-activated fibroblasts. By glycerol gradient ultracentrifugation, we also showed that: (a) in fibroblasts deficient in IKK, and IKK,, NEMO predominantly exists as a monomer; (b) in NEMO-deficient fibroblasts, IKK, dimers are present that are less stable than IKK, dimers. Intriguingly, in resting Rat-1 fibroblasts, 160-kDa IKK,,NEMO and IKK,,NEMO heterocomplexes were observed as well as a significant proportion of NEMO monomer. These results suggest that most NEMO molecules do not form a tripartite IKK complex with an IKK,,IKK, heterodimer as previously reported in the literature but, instead, NEMO is able to form a complex with the monomeric forms of IKK, and IKK,. [source] Conformational and functional analysis of the lipid binding protein Ag-NPA-1 from the parasitic nematode Ascaridia galliFEBS JOURNAL, Issue 1 2005Rositsa Jordanova Ag-NPA-1 (AgFABP), a 15 kDa lipid binding protein (LBP) from Ascaridia galli, is a member of the nematode polyprotein allergen/antigen (NPA) family. Spectroscopic analysis shows that Ag-NPA-1 is a highly ordered, ,-helical protein and that ligand binding slightly increases the ordered secondary structure content. The conserved, single Trp residue (Trp17) and three Tyr residues determine the fluorescence properties of Ag-NPA-1. Analysis of the efficiency of the energy transfer between these chromophores shows a high degree of Tyr-Trp dipole-dipole coupling. Binding of fatty acids and retinol was accompanied by enhancement of the Trp emission, which allowed calculation of the affinity constants of the binary complexes. The distance between the single Trp of Ag-NPA-1 and the fluorescent fatty acid analogue 11-[(5-dimethylaminonaphthalene-1- sulfonyl)amino]undecanoic acid (DAUDA) from the protein binding site is 1.41 nm as estimated by fluorescence resonance energy transfer. A chemical modification of the Cys residues of Ag-NPA-1 (Cys66 and Cys122) with the thiol reactive probes 5-({[(2-iodoacetyl)amino]ethyl}amino) naphthalene-1-sulfonic acid (IAEDANS) and N,N,-dimethyl- N -(iodoacetyl)- N,-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)ethylenediamine (IANBD), followed by MALDI-TOF analysis showed that only Cys66 was labeled. The observed similar affinities for fatty acids of the modified and native Ag-NPA-1 suggest that Cys66 is not a part of the protein binding pocket but is located close to it. Ag-NPA-1 is one of the most abundant proteins in A. galli and it is distributed extracellularly mainly as shown by immunohistology and immunogold electron microscopy. This suggests that Ag-NPA-1 plays an important role in the transport of fatty acids and retinoids. [source] Triplet Formation in Fullerene Multi-Adduct Blends for Organic Solar Cells and Its Influence on Device PerformanceADVANCED FUNCTIONAL MATERIALS, Issue 16 2010Clare Dyer-Smith Abstract In organic solar cells, high open circuit voltages may be obtained by choosing materials with a high offset between the donor highest occupied molecular orbital (HOMO) and acceptor lowest unoccupied molecular orbital (LUMO). However, increasing this energy offset can also lead to photophysical processes that compete with charge separation. In this paper the formation of triplet states is addressed in blends of polyfluorene polymers with a series of PCBM multi-adducts. Specifically, it is demonstrated that the formation of such triplets occurs when the offset energy between donor ionization potential and acceptor electron affinity is ,1.6 eV or greater. Spectroscopic measurements support a mechanism of resonance energy transfer for triplet formation, influenced by the energy levels of the materials, but also demonstrate that the competition between processes at the donor,acceptor interface is strongly influenced by morphology. [source] Constitutive oligomerization of human D2 dopamine receptors expressed in Spodoptera frugiperda 9 (Sf9) and in HEK293 cellsFEBS JOURNAL, Issue 19 2003Analysis using co-immunoprecipitation, time-resolved fluorescence resonance energy transfer Human D2Long (D2L) and D2Short (D2S) dopamine receptor isoforms were modified at their N-terminus by the addition of a human immunodeficiency virus (HIV) or a FLAG epitope tag. The receptors were then expressed in Spodoptera frugiperda 9 (Sf9) cells using the baculovirus system, and their oligomerization was investigated by means of co-immunoprecipitation and time-resolved fluorescence resonance energy transfer (FRET). [3H]Spiperone labelled D2 receptors in membranes prepared from Sf9 cells expressing epitope-tagged D2L or D2S receptors, with a pKd value of , 10. Co-immunoprecipitation using antibodies specific for the tags showed constitutive homo-oligomerization of D2L and D2S receptors in Sf9 cells. When the FLAG-tagged D2S and HIV-tagged D2L receptors were co-expressed, co-immunoprecipitation showed that the two isoforms can also form hetero-oligomers in Sf9 cells. Time-resolved FRET with europium and XL665-labelled antibodies was applied to whole Sf9 cells and to membranes from Sf9 cells expressing epitope-tagged D2 receptors. In both cases, constitutive homo-oligomers were revealed for D2L and D2S isoforms. Time-resolved FRET also revealed constitutive homo-oligomers in HEK293 cells expressing FLAG-tagged D2S receptors. The D2 receptor ligands dopamine, R -(,)propylnorapomorphine, and raclopride did not affect oligomerization of D2L and D2S in Sf9 and HEK293 cells. Human D2 dopamine receptors can therefore form constitutive oligomers in Sf9 cells and in HEK293 cells that can be detected by different approaches, and D2 oligomerization in these cells is not regulated by ligands. [source] Synthesis of Zwitterionic Water-Soluble Oligofluorenes with Good Light-Harvesting AbilityADVANCED FUNCTIONAL MATERIALS, Issue 13 2010Chengfen Xing Abstract A new water-soluble zwitterionic oligofluorene bearing carboxylic acid and quaternary ammonium as pendant groups (OF-1) is synthesized and characterized. It forms aggregates by intermolecular electrostatic interactions and exhibits similar light-harvesting ability as that of conjugated polymers. Efficient fluorescence resonance energy transfer (FRET) occurs from OF-1 to double-stranded DNA tagged with fluorescein (dsDNA-F1). A photoresponsive oligofluorene (OF-3) is also synthesized by protecting OF-1 with 1-(2-nitrophenyl)ethanol. Photolysis of OF-3 can produce OF-1 to result in a fluorescence "turn-on" response, thus the FRET from OF-3 to dsDNA-Fl can be turned on by light irradiation. OF-3 offers the potential for remote DNA sensing. [source] Bionanotechnology: Enhancement of Aggregation-Induced Emission in Dye-Encapsulating Polymeric Micelles for Bioimaging (Adv. Funct.ADVANCED FUNCTIONAL MATERIALS, Issue 9 2010Mater. Amphiphilic block copolymers can form polymer micelles for delivering hydrophobic fluorescent probes with aggregation-induced emission properties, as presented by A. K.-Y. Jen et al. on page 1413. By itself, 1,1,2,3,4,5-hexaphenylsilole (HPS) exhibits dramatically enhanced blue-green fluorescent emission efficiencies when encapsulated within the hydrophobic core of a polymeric micelle. When HPS is co-encapsulated with bis(4-(N -(1-naphthyl) phenylamino)-phenyl)fumaronitrile, effective orange-red fluorescence resonance energy transfer can be demonstrated within live RAW 264.7 cells. Illustration provided by Brent Polishak. [source] Enhancement of Aggregation-Induced Emission in Dye-Encapsulating Polymeric Micelles for BioimagingADVANCED FUNCTIONAL MATERIALS, Issue 9 2010Wen-Chung Wu Abstract Three amphiphilic block copolymers are employed to form polymeric micelles and function as nanocarriers to disperse hydrophobic aggregation-induced emission (AIE) dyes, 1,1,2,3,4,5-hexaphenylsilole (HPS) and/or bis(4-(N -(1-naphthyl) phenylamino)-phenyl)fumaronitrile (NPAFN), into aqueous solution for biological studies. Compared to their virtually non-emissive properties in organic solutions, the fluorescence intensity of these AIE dyes has increased significantly due to the spatial confinement that restricts intramolecular rotation of these dyes and their better compatibility in the hydrophobic core of polymeric micelles. The effect of the chemical structure of micelle cores on the photophysical properties of AIE dyes are investigated, and the fluorescence resonance energy transfer (FRET) from the green-emitting donor (HPS) to the red-emitting acceptor (NPAFN) is explored by co-encapsulating this FRET pair in the same micelle core. The highest fluorescence quantum yield (,62%) could be achieved by encapsulating HPS aggregates in the micelles. Efficient energy transfer (>99%) and high amplification of emission (as high as 8 times) from the NPAFN acceptor could also be achieved by spatially confining the HPS/NPAFN FRET pair in the hydrophobic core of polymeric micelles. These micelles could be successfully internalized into the RAW 264.7 cells to demonstrate high-quality fluorescent images and cell viability due to improved quantum yield and reduced cytotoxicity. [source] Nanoparticle Electroluminescence: Controlling Emission Color Through Förster Resonance Energy Transfer in Hybrid ParticlesADVANCED FUNCTIONAL MATERIALS, Issue 22 2009Christopher F. Huebner Abstract Electroluminescent (EL) polymers are attractive for developing all-organic light-emitting devices (OLEDs) due to the potential advantages that polymeric systems may offer in the large-scale manufacturing of electronics. Nonetheless, many of these EL , -conjugated polymers are inherently insoluble in the solvents employed in the intended solution-based manufacturing processes. One such polymer is poly(2,5-dioctyl-1,4-phenylenevinylene) (POPPV), where the inherent lack of solubility of POPPV in organic solvents has frustrated its widespread application in devices and no OLEDs have been presented that exploit its electroluminescence characteristics. In this effort, a unique strategy is presented for the preparation of hybrid nanoparticles composed of POPPV, a green emitter (,em,=,505,nm) and poly(9,9-di- n -octylfluorenyl-2,7-diyl) (PFO), a blue emitter (,em,=,417,nm). The aqueous-based nanoparticle dispersion composed of these hybrid particles is stable to aggregate and can be employed in the construction of OLEDs. The color characteristics of the electroluminescence for the devices can be tuned by exploiting the Förster resonance energy transfer between the polymers within a particle, while suppressing energy transfer between the particles. These aqueous-based nanoparticle dispersions are amenable to being printed into devices through high-throughput manufacturing techniques, for example, roll-to-roll printing. [source] Simple and Efficient Generation of White Light Emission From Organophosphorus Building BlocksADVANCED FUNCTIONAL MATERIALS, Issue 22 2009Carlos Romero-Nieto Abstract This paper describes a structure,property study using two dithieno[3,2- b;2,,3,- d]phosphole building blocks for the generation of white light emission and the incorporation of these units in a single polystyrene material. The emission of one of the light-emitting organophosphorus building blocks can efficiently be switched from orange to green by simple protonation of the amino functional groups that are part of the , -conjugated scaffold. The resulting three components (blue, green, and orange) exhibit photophysical properties that allow for an efficient fluorescence resonance energy transfer (FRET) in the mixture/polymer and provide intense white fluorescence upon excitation of the blue component; the fluorescence is close to pure white in solution and similar to the emission of an incandescent light bulb in the thin film. The results nicely illustrate the intriguing features that can be obtained by exclusively using organophosphorus-based organic electronic materials. [source] Conjugated Polymers Combined with a Molecular Beacon for Label-Free and Self-Signal-Amplifying DNA MicroarraysADVANCED FUNCTIONAL MATERIALS, Issue 20 2009Kangwon Lee Abstract A conjugated polymer (CP) and molecular-beacon-based solid-state DNA sensing system is developed to achieve sensitive, label-free detection. A novel conjugated poly(oxadiazole) derivative exhibiting amine and thiol functional groups (POX-SH) is developed for unique chemical and photochemical stability and convenient solid-state on-chip DNA synthesis. POX-SH is soluble in most nonpolar organic solvents and exhibits intense blue fluorescence. POX-SH is covalently immobilized onto a maleimido-functionalized glass slide by means of its thiol group. Molecular beacons having a fluorescent dye or quencher molecule as the fluorescence resonance energy transfer (FRET) acceptor are synthesized on the immobilized POX-SH layer through direct on-chip oligonucleotide synthesis using the amine side chain of POX-SH. Selective hybridization of the molecular beacon probes with the target DNA sequence opens up the molecular beacon probes and affects the FRET between POX-SH and the dye or quencher, producing a sensitive and label-free fluorescence sensory signal. Various molecular design parameters, such as the size of the stem and loop of the molecular beacon, the choice of dye, and the number of quencher molecules are systematically controlled, and their effects on the sensitivity and selectivity are investigated. [source] Spectral imaging fluorescence microscopyGENES TO CELLS, Issue 9 2002Tokuko Haraguchi The spectral resolution of fluorescence microscope images in living cells is achieved by using a confocal laser scanning microscope equipped with grating optics. This capability of temporal and spectral resolution is especially useful for detecting spectral changes of a fluorescent dye; for example, those associated with fluorescence resonance energy transfer (FRET). Using the spectral imaging fluorescence microscope system, it is also possible to resolve emitted signals from fluorescent dyes that have spectra largely overlapping with each other, such as fluorescein isothiocyanate (FITC) and green fluorescent protein (GFP). [source] Controlling Affinity Binding with Peptide-Functionalized Poly(ethylene glycol) HydrogelsADVANCED FUNCTIONAL MATERIALS, Issue 14 2009Chien-Chi Lin Abstract Poly(ethylene glycol) (PEG) hydrogels functionalized with peptide moieties have been widely used in regenerative medicine applications. While many studies have suggested the importance of affinity binding within PEG hydrogels, the relationships between the structures of the peptide motifs and their binding to protein therapeutics remain largely unexplored, especially in the recently developed thiol-acrylate photopolymerization systems. Herein, Förster resonance energy transfer (FRET) and thiol-acrylate photopolymerizations are employed to investigate how the architectures of affinity peptides in crosslinked hydrogels affect their binding to diffusible proteins. The binding between diffusible streptavidin and biotinylated peptide immobilized to PEG hydrogel network was used as a model system to reveal the interplay between affinity binding and peptide sequences/architectures. In addition, peptides with different structures are designed to enhance affinity binding within PEG hydrogels and to provide tunable affinity-based controlled delivery of basic fibroblast growth factor (bFGF). This study demonstrates the importance of affinity binding in controlling the availability of hydrogel-encapsulated proteins and provides strategies for enhancing affinity binding of protein therapeutics to bound peptide moieties in thiol-acrylate photopolymerized PEG hydrogels. The results presented herein should be useful to the design and fabrication of hydrogels that retain and exhibit sustained release of growth factors for promoting tissue regeneration. [source] Intercalating Dye Harnessed Cationic Conjugated Polymer for Real-Time Naked-Eye Recognition of Double-Stranded DNA in SerumADVANCED FUNCTIONAL MATERIALS, Issue 9 2009Kan-Yi Pu Abstract Thiazole orange (TO), an intercalating dye, is integrated into cationic poly(fluorene- alt -phenylene) (PFP) to develop a macromolecular multicolor probe (PFPTO) for double-stranded DNA (dsDNA) detection. This polymer design not only takes advantage of the high affinity between TO and dsDNA to realize dsDNA recognition in biological media, but also brings into play the light-harvesting feature of conjugated polymers to amplify the signal output of TO in situ. PFPTO differentiates dsDNA from single-stranded DNA (ssDNA) more effectively upon excitation of the conjugated backbone relative to that upon direct excitation of TO as a result of efficient fluorescence resonance energy transfer from the polymer backbone to the intercalated TO. In the presence of dsDNA, energy transfer within PFPTO is more efficient as compared to that for free TO/PFP system, which leads to better dsDNA discriminability for PFPTO in contrast to that for TO/PFP. The distinguishable fluorescent color for PFPTO solutions in the presence of dsDNA allows naked-eye detection of dsDNA with the assistance of a hand-held UV lamp. The significant advantage of this macromolecular fluorescent probe is that naked-eye detection of label-free dsDNA can be performed in biological media in real-time. [source] A Smart Nanoprobe Based On Fluorescence-Quenching PEGylated Nanogels Containing Gold Nanoparticles for Monitoring the Response to Cancer TherapyADVANCED FUNCTIONAL MATERIALS, Issue 6 2009Motoi Oishi Abstract A biocompatible, caspase-3-responsive, and fluorescence-quenching smart apoptosis nanoprobe based on a PEGylated nanogel that contains gold nanoparticles (GNPs) (fluorescence quenchers) in the cross-linked polyamine gel core and fluorescein isothiocyanate (FITC)-labeled DEVD peptides at the tethered PEG chain ends is prepared for monitoring the cancer response to therapy. FITC,DEVD,nanogel,GNP shows very little fluorescence in the absence of activated caspase-3 (normal cells) through the fluorescence resonance energy transfer (FRET) process between the GNPs and the FITC molecules, while pronounced fluorescence signals are observed in apoptotic cells because of the cleavage of the DEVD peptide by activated caspase-3 present in the cells, which results in the release of FITC molecules. Thus, remarkable quenching and dequenching of fluorescence signals in response to activated caspase-3 is observed. Apoptotic cells are detected in human hepatocyte (HuH-7) multicellular tumor spheroids (MCTSs), a commonly used three-dimensional in vitro model mimicking the in vivo biology of tumors, as early as one day post-treatment with staurosporine, an apoptosis-inducing agent; while growth inhibition (i.e., change in size) of the HuH-7 MCTSs is only observed after a delay of three days (i.e., on day 4). This demonstrates the effectiveness of the FITC,DEVD,nanogel,GNP probe as a smart nanoprobe for real-time monitoring as well as a more rapid assessment of the early response to cancer therapy. [source] | ||||||||||||||||||||||||||||||||||